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Bacterial Urease and its Role in Long-Lasting Human Diseases

Bacterial Urease and its Role in Long-Lasting Human Diseases Send Orders of Reprints at bspsaif@emirates.net.ae Current Protein and Peptide Science, 2012, 13, 789-806 789 1,* 1 1 2 Iwona Konieczna , Paulina  arnowiec , Marek Kwinkowski , Beata Kolesi ska , 2 2 1 Justyna Fr czyk , Zbigniew Kami ski and Wies aw Kaca Department of Microbiology, Institute of Biology, The Jan Kochanowski University, ul. wi tokrzyska 15, 25-406 Kielce, Poland; Technical University of Lodz, Institute of Organic Chemistry, ul. eromskiego 116, 90-924  ód , Poland. Abstract: Urease is a virulence factor found in various pathogenic bacteria. It is essential in colonization of a host organ- ism and in maintenance of bacterial cells in tissues. Due to its enzymatic activity, urease has a toxic effect on human cells. The presence of ureolytic activity is an important marker of a number of bacterial infections. Urease is also an immuno- genic protein and is recognized by antibodies present in human sera. The presence of such antibodies is connected with progress of several long-lasting diseases, like rheumatoid arthritis, atherosclerosis or urinary tract infections. In bacterial ureases, motives with a sequence and/or structure similar to human proteins may occur. This phenomenon, known as mo- lecular mimicry, leads to the appearance of autoantibodies, which take part in host molecules destruction. Detection of an- tibodies-binding motives (epitopes) in bacterial proteins is a complex process. However, organic chemistry tools, such as synthetic peptide libraries, are helpful in both, epitope mapping as well as in serologic investigations. In this review, we present a synthetic report on a molecular organization of bacterial ureases - genetic as well as structural. We characterize methods used in detecting urease and ureolytic activity, including techniques applied in disease diagnos- tic processes and in chemical synthesis of urease epitopes. The review also provides a summary of knowledge about a toxic effect of bacterial ureases on human body and about occurrence of anti-urease antibodies in long-lasting diseases. Keywords: Antibodies, long-lasting diseases, synthetic peptides, urease 1. INTRODUCTION in long-lasting diseases, like atherosclerosis or rheumatoid arthritis [6, 7]. This phenomenon will be precisely described Urease (urea amidohydrolase; EC 3.5.1.5) was the first in the next paragraphs. enzyme to be crystallized (1926). It was also the first enzy- matic protein in which the presence of nickel ions was noted 2. OCCURRENCE OF UREASE PRODUCING OR- [1]. Since then, an intensive study on urease has been con- GANISMS ducted, thanks to which a role of urease in nitrogen com- pounds circulation has been determined. It has also been Urease is produced by many different bacteria [8-17], showed that urease may be a virulence factor essential in fungi [3, 18, 19], plants [1, 3, 8, 20, 21] and even some in- various illnesses, including long-lasting diseases. vertebrates [20, 21]. Microorganisms with ureolytic proper- Urease is capable of urea hydrolysis. This compound is ties were found in soil and water as well as in human and widespread: it is found in the natural environment (water and animal bodies [8]. Ureolytic bacteria may belong to symbi- soil) and in human body, where its occurrence is connected otic natural microflora or to pathogens. In facultative anaer- with protein degradation. In humans, urea is a factor of nor- obes from intestinal microflora the level of this activity is mal functions of kidneys [2, 3]. A healthy adult excretes diverse and species characteristic [9]. about 30 g of urea per day [2]. However, it is present not Ureolytic activity is often observed in pathogenic bacte- only in urine, but also in blood serum, sweat and even in ria. Such a feature is characteristic of pathogenic Staphylo- stomach [1, 2]. Hydrolysis of urea by urease is a complex coccus strains. Over 90% of clinical methicillin resistant process. In the first step, one molecule of ammonia and one Staphylococcus aureus strains are capable of urea hydrolysis molecule of carbamate appear. In water solution, carbamate [10]. Staphylococcus leei isolated from biopsy material from spontaneously converts into the second ammonia molecule gastritis patients was also ureolytic. Uropathogenic Staphy- and carbonic acid. Next ammonia is protonated (Fig. 1). This lococcus saprophiticus is also capable of this activity [1, 11, process results in pH increase [1]. 22]. Urease is observed in Helicobacter sp., including all Urease and ammonia, generated during urea hydrolysis, Helicobacter pylori isolated from gastritis patients [1, 4, 23]. may be toxic for human tissue [4, 5] and probably have role Urease is an enzyme synthesized by pathogenic mycobacte- ria like Mycobacterium tuberculosis and Mycobacterium *Address correspondence to this author at the Department of Microbiology, bovis [12]. It was observed that anaerobic clostridia are ca- Institute of Biology, The Jan Kochanowski University, ul. wi tokrzyska pable of urea hydrolysis. About 2% of Clostridium perfring- 15, 25-406 Kielce, Poland; Tel: 48 41 349 63 05; Fax: 48 41 349 62 92; ers strains, an etiologic factor of gas gangrene, showed this E-mail: iwona.konieczna@ujk.edu.pl 1389-2037/12 $58.00+.00 © 2012 Bentham Science Publishers 790 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (1). Scheme of urea hydrolysis. feature [13]. Even some strains of Vibrio parahaemolyticus, environmental bacteria. Unique urease of Helicobacter sp. a species considered non-ureolytic, produce urease [14]. An- has a different structure. In H. pylori, urease consists of only other generally urease negative bacterial species is Es- two subunits: 26.5 kDa UreA (subunit  ) and 61.7 kDa UreB cherichia coli. Among E. coli strains, about 1% of urease- (subunit  ) coded by ureA and ureB genes [29]. A smaller positive isolates were found. This feature was connected Helicobacter sp. urease structural gene (ureA) corresponds with pathogenic O111, O157:H7, O145 and O26 enterohem- with a hypothetical fusion gene arisen from ureA and ureB orrhagic E. coli, and in O157 serogroup with sorbitol fer- typical of other bacteria, while a larger gene (ureB) is analo- menting, but non motile strains [15-17]. Proteus mirabilis is gous to ureC (Fig. 2) [30-33]. a well-known ureolytic human’s pathogen. Urease is one of Urease composed of two different polypeptides (21 kDa the major bacterial virulence factors during urinary tract in- and 65 kDa) was also identified in SL100 ureolytic coccoid fections caused by these bacteria [1, 24]. A similar phe- strain isolated from stomach biopsy material. This strain was nomenon was noted for uropathogenic Ureapasma urealyti- related to Staphylococcus cohnii and Staphylococcus xylosus, cum, Klebsiella spp., Pseudomonas spp., Corynebacterium which possess three urease subunits [34]. sp. D2, Proteus penneri, Providencia stuartii and Morgan- An active center of enzyme with two metal ions is lo- ella morganii [1, 22]. cated in the largest of structural subunits. In all ureases it is designed as UreC, except Helicobacter sp., in which case it 3. GENETIC AND STRUCTURAL ORGANIZATION is UreB [1]. Ureases are nickel-containing enzymes; how- OF BACTERIAL UREASES ever, for microaerophilic Helicobacter mustelae an iron- Urease is a nickel-containing enzyme, which requires containing urease was revealed [23]. activity of a few additional proteins for acquisition of its All bacterial ureases occur as inactive apoenzymes com- hydrolytic properties. This process involves genes coding posed of three or two types of polypeptides coded by specific structural enzyme polypeptides as well as genes coding ac- structural genes. However, additional proteins, products of cessory proteins, located in a joint cluster [1, 25]. accessory genes are required for urease activation. Those Bacterial ureases are always multimeric enzymes com- proteins (UreD, UreE, UreF, UreG and UreH) are involved posed of two or three different polypeptides [1]. In P. in transporting nickel ions into a cell and in incorporating mirabilis, three structural subunits: 11 kDa UreA (subunit  ), them into an active center of apoenzyme [35-44]. P. mirabi- 12.2 kDa UreB (subunit  ) and 61 kDa UreC (subunit  ) are lis produce active urease in presence of urea. In these bacte- found [1, 26, 27]. These polypeptides are encoded by three ria a regulatory gene ureR is present (see Fig. 2). Its product structural genes: ureA, ureB and ureC respectively [28]. is a urea inducible regulator controlling expression of re- Such organization is characteristic of most pathogenic and maining genes [1]. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 791 Fig. (2). Scheme of genetic organization of urease genes and structural composition of urease. Genetic organization of ure genes was performed basing on Microbial Genome Viever MGV 2.0 (http://mgv2.cmbi.ru.nl) for H. pylori G27, P. mirabilis HI4320, K. pneumonia 342 and Yersinia enterocolitica 8081. Under genes/polypeptides are sizes of particular genes as well as a number of amino acids of particular polypeptide were taken from NCBI database for records: CP001173 (H. pylori G27), AM942759 (P. mirabilis HI4320), NC_011283 (K. pneumonia 342) and NC_008800 (Y. enterocolitica 8081); structural genes as well as urease subunits are underlined. A highly mobile helix-turn-helix motif, located in  remaining polypeptides, similarities were smaller (Table 1.) subunit and called “flap” is essential for urease activity (see [47, 48]. Fig. 5). It may adopt two different conformations. In the In  subunit, the active center is the most conservative. “open” position, urea may enter into the active site, where Particularly stable are nickel ligands: histidines (in K. aero- hydrolyze is performed. In the “closed” position, flap covers genes His-134, His-136, His-246 and His-272), lysine (in K. the active center and blocks access to it [25]. aerogenes Lys-217) and aspartic acid (in K. aerogenes Asp- Active ureases are heterooligomeric complexes. How- 360) [1, 51]. Similarities were also observed for the flap ever, the number of particular structural subunits is always fragment from over 160 sequences of  subunit of ureases equal. In K. aerogenes urease, as well as in other tree-subunit from different microorganisms, including human pathogens. bacterial ureases, UreC/UreB/UreA molecules occur in the Many sequential identities occurred in all amino acid se- ratio 1:1:1. Likewise, for Helicobacter sp. UreB/UreA are quences. In bacteria causing human diseases, even not always in the ratio 1:1 [1]. closely related, significant conservatism was noted (Fig. 4) [52]. Urease from K. aerogenes, as well as the most of other with three active centers, one The structure of a flap region in ureases from different bacteria, is triple trimer () bacteria also possess similar conformation (Fig. 5). Bacterial in each of three  subunits. Amino- and carboxyl terminus of pathogens shows different ureolytic activity. Methods for its each subunit are free and they are able to bind additional detection, including techniques applied in disease diagnosis, compounds without disturbing the enzyme structure [1]. But are diverse. Prochrorococcus marinus sp. PCC 9511 produces urease composed according to () pattern [1, 45]. Enzymes from 5. METHODS FOR DETECTING OF UREOLYTIC Helicobacter sp. may form a more complex structure, built ACTIVITY from 12 subunits. Polypeptides  and  are linked forming trimer ( ) , where N-terminal domain of  subunit are es- Hydrolysis of urea is one of useful features in the bacte- sential in aggregation process. Then, four such trimers form ria identification. In a few infections caused by microbes, a tetrahedral complex (Fig. 3). detection of this activity is essential to disease diagnosis. Probably, such a highly complex structure of H. pylori Numerous assays are available to determine urease activ- urease enables its activity in acidic conditions, when other ity as well as to analyze kinetic behavior of urease. Most of ureases undergo nonreversible inactivation [46]. them are indirect and based on colorimetric detection of ammonia released during incubation with a buffered urea 4. CONSERVATISM OF BACTERIAL UREASES solution [53]. Ureases are considered to be conservative enzymes. One of the first methods was detecting bacterial ureolytic Among all Ure polypeptides from different bacterial species, activity based on the cultivation of microorganism on urea the highest sequence similarities were observed between containing medium (Christensen's urea medium) [54]. This is structural urease subunits from different sources. In case of the most popular qualitative method using for uropathogenes 792 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (3). Scheme of structure of bacterial ureases. Domain organization is reported for (A) H. pylori and (B) Bacillus pasteurii urease mono- mers, (C) B. pasteurii and (D) H. pylori urease trimers, and (E) H. pylori dodecamer. Table 1. The Sequence Similarity of Structural and Accessory Polypeptides of Ureases of Different Bacteria. Polypeptides Identical amino acid sequence [%] Reference Bordetella Alcaligenes K. aerogenes P. mirabilis bronchiseptica eutrophus (UreA) 84 79 73 (UreB) 63 69 67 (UreC) 70 69 68 [49] UreD 43 33 28 UreG 75 66 59 UreE 39 38 38 UreF 54 31 31 Actinobacillus Haemophilus Bacillus sp. H. pylori K. aerogenes pleuropneumoniae influenzae TB-90 (UreA) 96 67 64 57 (UreB) 86 55 53 60 (UreC) 87 62 62 66 [50] UreE 85 21 28 25 UreF 67 34 34 27 UreG 95 60 66 60 UreD 70* 24 29* 21 * - data for UreH Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 793 Fig. (4). Alignment of the amino acid sequence of urease flap fragment from pathogenic bacteria. Black - amino acid present in at least 80% of compared sequences, grey - amino acid present in at least 70% of compared sequences, white - amino acid present in less than 70% of compared sequences. Sequences from NCBI database (Accession Numbers are in figure). Alignment was performed by Clustal W 2.1 and edited with GeneDoc. Fig. (5). Conformational conservatism of bacterial ureases. All models of bacterial ureases were from ExPASy SIB Bioinformatics Resource Portal (Q7X3W5 - H. pylori; P16122 - P. hauserii; Q6GEE4 - S. aureus); overlapping was performed with RasWin Molecular Graphics Visualisation Tool (http://rasmol.org/). A - structure of H. pylori (blue) and P. hauserii (white) ureases, B - structure of H. pylori (blue) and S. aureus (white) ureases; flap region is marked by a white ellipse. like Proteus sp. In case of this bacterial species, results may most popular invasive test is a rapid urease test (RUT) that be obtained even after 4 h. A modification of Christensen requires obtaining tissue samples. However, this method is technique allows reducing assay time [54, 55]. inconvenient for patients and also incurs high costs [57, 58]. It requires biopsy specimens from defined regions of the Ureolytic activity is one of biomarkers employed to di- stomach. This material is placed on a urea-containing me- agnose H. pylori infection and to monitor bacteria eradica- dium. If bacteria are present in the specimen, the change of tion by drug treatment [56]. color resulted from alkalization of the medium is observed For diagnosis, invasive and noninvasive tests, depending [59]. A urea breath test (UBT) is commonly used among on whether endoscopy is required or not, are applied. The noninvasive tests. This method is simple, but its performance 794 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. may be slightly complicated in case of very young children urea. This property may be due to mobility of the flap region, as well as patients with certain neurological disorders [49, 50 which is different than in K. aerogenes or B. pasteurii 60]. It involves oral administration of a nontoxic isotopically ureases [46]. Due to the high activity of H. pylori urease, 14 13 labeled (C or C ) urea to a patient. Urea is hydrolyzed by local microenvironment surrounding bacterium becomes H. pylori to ammonia and isotope-containing CO . Carbon nearly neutral. Moreover, live bacterial cells adsorb on the dioxide is dissolved into blood and removed via lungs. Iso- surface enzymes released upon other H. pylori autolysis, topes are detected in exhaled air. This is a test of choice in which makes it possible for them to get to gastric mucus medical practice for detecting H. pylori infection. [61]. There layer safely [80]. Ureolytic activity is essential for surviving are also suggestions that a urea breath test may be applied for M. tuberculosis, an etiologic factor of tuberculosis, a long- diagnosing tuberculosis [62]. lasting inflammatory lung disease. Bacteria infect macro- phages. They reside in phagosome, where alkalization due to Other methods are used mainly in scientific research. ureolytic activity and subvert phagosome maturation takes Each of the methods developed for determination of ure- place. Additionally, urease activity enables bacterium to ex- olytic activity has some advantages and disadvantages (Table ist in the environment where nitrogen sources are limited to 2). urea [81]. Ureolytic activity is useful in better surviving of There are a lot of other methods based on detecting am- bacteria also in case of uropathogenes. Urease facilitates monia released by urease action, which can be determined by urinary tract infection. Infection dose for ureolytic P. mirabi- vacuum distillation, a microdiffusion, steam distillation and lis HI4320 was 1000-times lower in comparison with its electroconductivity measurement [77]. non-ureolytic mutant. Urease activity raises pH of human urine, which allows precipitation of normally soluble polyva- Fourier Transform Infrared (FTIR) spectroscopy is a lent ions to struvite and carbonate apatite. These compounds method raising a big hope for easy, quick and continuous aggregate around bacteria, forming urinary stones. Inside detection of ureolytic activity. This technique constitutes a such stones, microorganisms are protected from antibiotics radically different approach to enzymatic activity determina- and the host’s immune system [24, 83]. Urinary stones block tion. It is based on the measurement of molecular vibrations urethra or catheters leading to acute bacteriuria [24]. The energy of functional groups in organic compounds. This role of ureolytic activity in urinary stones formation was also makes FTIR spectroscopy a highly sensitive and reproduci- showed for U. urealyticum, S. saprophiticus, S. aureus and ble method. Unlike the previously discussed methods, this some Klebsiella spp., Pseudomonas spp., as well as Coryne- technique enables continuous monitoring of enzymatic reac- bacterium sp. D2, P. penneri, P. stuartii, M. morganii [1, tion by a simultaneous analysis of disappearance of substrate 22]. and the appearance of product. However, substrate as well as product must have different spectra. FTIR spectroscopy also One of the features essential in bacterial infections is enables enzyme kinetics investigation [78]. Attenuated Total persistence to the host’s cells. Schoep et al. showed that H. Reflection Fourier Transform Infrared (ATR-FTIR) spec- pylori urease have two sites (one at the N-termini of UreA troscopy was recommended to enzymatic activity analysis subunit and the other at C-termini of UreB) which were in- [79]. This technique was also applied for urease activity in- volved in persistence to endothelial cells during mouse colo- vestigations. Bands of absorbance characteristic of urea nization [29]. This observation was confirmed by investiga- (substrate) and of bicarbonate (product) could be easily tions with urease-negative H. pylori mutants incapable of monitored in time intervals (Fig. 6). [Zarnowiec et al., data colonization [1,4]. Moreover, also urease released from unpublished]. However, ATR-FTIR spectroscopy is now lyzed bacterial cells is capable of adsorption into the mucus used only in research applications, not in routine medical layer [4]. practice. Bacterial ureases affect host immune system cells. In H. pylori infection, this metalloenzyme activates monocytes and 6. UREASE AS A PATHOGENIC BACTERIA VIRU- neutrophils, which leads to secretion of inflammatory cyto- LENCE FACTOR kines and causes indirect damage to epithelial cells. Urease Bacterial ureases play a role in disease pathogenesis. is a chemotactic factor for monocytes and neutrophils. In- They are connected with urinary stones occurrence and flammatory reaction may also be initiated by adsorption of catheters blocking, pyelonephritis, ammonia encephalopathy, released enzyme into the mucus layer [4]. Induction of in- hepatic coma as well as gastritis. In many papers there are flammatory reaction was also observed for Y. enterocolitica information concerning toxic effects of bacterial ureases urease. Ability of bacterial UreB subunit to induce experi- (Table 3). mental reactive arthritis was revealed [1, 84]. The role of urease in bacterium surviving in unfavorable Urease may contribute in damaging host’s cells. Enzyme microenvironment in the host’s body is especially noticeable from H. pylori stimulates expression of inducible NO- in case of H. pylori, a causative agent of gastritis and peptic synthesizing enzyme (iNOS), which may have a cytotoxic ulceration [1, 4]. At in vitro conditions, H. pylori is sensitive effect [80]. Urease may exert a toxic effect also indirectly, to low pH. During infection, microorganisms have to pass by ammonia - the product of urea hydrolysis. During H. py- through gastric acid before reaching the protective mucus lori infection, a stimulation of an oxidative burst in neutro- layer. In these circumstances, a pathogen produces a large phils ensues and there is a release of hydrogen peroxide, amount of urease which is not observed in other bacteria which next oxidizes chlorine ions. Ammonia generated by [80]. At low pH, enzymatic activity of H. pylori urease is urease reacts with them and gives toxic monochloramine [1]. probably connected with its dodecameric structure. This en- Johnson et al. revealed, using mouse model, that ammonia zyme is also able to perform a more efficient hydrolysis of causes tissue damage also during urinary tract infections Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 795 Table 2. Characteristic of Methods for Ureolytic Activity Determination. Method Description Advantages Disadvantages Application Reference Qualitative activity is detected on the microbiological medium restricted to cultivable bacte- containing urea and phenol ria able to grow on this me- suitable for routine detection Urea-phenol facility of realization [53] red as pH indicator. Bacteria dium of activity, not recommended red-agar plate inexpensive [63] for kinetic analysis alkalize medium by hydroly- results need multiplication of sis of urea, causing change bacteria of its color. pH-dependent method. Sam- ple containing urease is electrophoresed in native allows estimation of the size equipment for electrophore- Native gel agarose or acrylamide gel. [64] of active protein sis is indispensable electrophoresis Active protein is detected [65] inexpensive time consuming after incubation of gel in solution containing urea and phenol red. Quantitative requires numerous sampling spectrophotometric method of the reaction mixture based on detection of ammo- simple [8] Phenol – sensitive to various factors very useful in full kinetic nia released during urea able to detect even a small [66] hypochlorite like temperature and time, analyses, the most frequently hydrolysis. Ammonia reacts amount of ammonia (<0.02 [67] assay pH of buffers, inhibitors used in scientific research with phenol-hypochlorite at  mol) [68] limited linearity of the cali- high pH forming indophenol bration plots needs titration with diluted HCl to determine ammonia spectrophotometric assay Nesslerization amount [8] with Nessler reagent in col- easy to perform reaction long reaction time [69] ored pH indicator solution less sensitive than phenol- hypochlorite assay spectrophotometric method GLDH has pH optimum based on coupling reaction sensitive higher than most ureases [70] Coupled of ammonia with  - alternatively, a horseradish sensitive to inhibitors [71] enzyme assay ketoglutarate in presence of peroxidase may be used for [72] difficult interpretation glutamate dehydrogenase ammonia detection expensive (GLDH) interference by potassium and other monovalent ions method of direct monitoring unaffected by inhibitors low sensitive (in ion- useful in determination of Potentiometric of ammonia ions with ion- fast in performance [73] selective electrode) the urease inhibition mecha- assays [74] selective electrodes or am- allow continuous monitoring during assay, an ionic nisms monia-selective electrode of activity strength of solution changes (there is no buffer) methods based on the urea with carbon isotope radioac- need scintillation counter useful in diagnosis of H. Isotopic [75] tive C or non-radioactive fast in performance (for C ) or mass spectrome- pylori gastric mucosa infec- methods C (there are also methods [76] 13 15 ter (for C and N ) tion based on N ). A isotope- labeled CO is detected with ureolytic P. mirabilis. In kidneys, an acute inflamma- visible [87]. Moreover, ammonia released by urease causes tion as well as necrotic cells were observed. After one week, damage to the glycosaminoglycan layer in urothelial surface, pyelonephritis was in progress. Struvite stones were noted. and disturbs its protective function [5]. After two weeks, kidneys were ulcerated and fibrosis was 796 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (6). ATR FTIR spectra of the substrate (urea), product (NaHCO ), in reaction mixture containing 0.4  g urease from Canavalia ensi- formis. Spectra of the reaction mixture were recorded at several time intervals: 0, 10, 20 and 35 min, as indicated. Table 3. Pathologic Effect of Bacterial Ureases in Human Diseases. Role of urease Bacterium species Disease Reference H. pylori gastritis, peptic ulcers [1, 4, 80] Surviving in host’s organism M. tuberculosis tuberculosis [81] E. coli hemorrhagic colitis, HUS [82] Persistence to host’s cells H. pylori gastritis, peptic ulcers [29] P. mirabilis, M. morganii, U. urealyticum Precipitation of polyvalent ions urinary tract infections [1, 22, 24, 83] and others H. pylori gastritis, peptic ulcers [4] Stimulation of inflammatory reaction Y. enterocolitica reactive arthritis [1, 84] Cytotoxic effect on host’s cells H. pylori gastritis, peptic ulcers [1, 80] Damage to glycosaminoglycan layer P. mirabilis urinary tract infections [5] Damage of tight junctions H. pylori peptic ulcers [85] Aggregation of blood platelets H. pylori gastritis, cardiovascular disease [86] HUS - hemolytic uremic syndrome Recently, a new role of H. pylori urease has been estab- lipoxygenase-mediated pathway. Such properties may have a lished. During an infection, bacteria cause increased phos- role not only in gastrointestinal, but also cardiovascular dis- phorylation of the myosin regulatory light chain. Such phos- eases [86]. phorylation regulates the function of epithelial tight junction 7. PRESENCE OF ANTI-UREASE ANTIBODIES IN complexes, which have a role in maintenance of barrier func- SERA OF PATIENTS WITH LONG-LASTING DIS- tion, cell polarity as well as intercellular adhesion. Disrup- EASES tion of tight junction is associated with a carcinogenesis process. Wroblewski et al. showed that H. pylori urease may Bacterial ureases are considered to be one of the major be connected with gastric cancer by causing damage to tight antigens in several human diseases [1, 83, 84, 88]. Hirota et junctions [85]. al. showed that this protein is immunogenic [89]. In the flap region of enzyme, the ELR motive associated with immuno- Lately, a mechanism of activating blood platelets by bac- genic antigens is present (see Fig. 4) [90]. In long-lasting terial urease has been described. Wassermann et al. showed diseases caused by ureolytic bacteria, urease may stimulate that H. pylori enzyme stimulates this process through a generation of antibodies. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 797 Infections of H. pylori are mostly chronic and, in many region of urease react with bacterial antigen. However, they cases, lifelong [91]. During a infection, an elevated level of may also recognize IKEDV motive in CCRL1 (due molecu- immunoglobulins (secretory as well as circulating) was ob- lar mimicry) and cause an inflammatory process (Fig. 7) [6]. served [88]. Different classes of antibodies were noted: in Rheumatoid arthritis (RA) is a classic long-lasting dis- the stomach - IgA and IgM, in serum - IgG and IgA. IgG ease. It is an inflammatory condition leading to joint injury. immunoglobulins remain even for a few months after bacte- During its progress, hyaline cartilages of joints as well as rium eradication [92]. Urease from this bacterium is one of bones undergo atrophy [98]. Etiology of RA is complex and, the major immunodominant antigens [93]. It is considered a despite many years of investigations, still unclear. Apart vaccine in preventing H. pylori infections. In animal model, from genetic background of RA occurrence, a role of infec- vaccination with H. pylori urease provides a significant and tious agents, like P. mirabilis, Borrelia burgdorferi, Myco- long term protection against a bacterial infection. In humans, plasma sp., M. tuberculosis, E. coli, and Porphyromonas oral administration of such a vaccine resulted in a strong gingivalis as well as some viruses was discussed [99, 100]. immune response with minimal side effects [88]. Some of them are capable of urease synthesis [12, 24]. The presence of anti-urease antibodies in H. pylori sero- Among them, the most important is P. mirabilis. Wilson et positive individuals is correlated with age and living in al. revealed a connection of bacterial urease with disease highly developed regions. Leal-Herrera, in the investigations progress. They showed a molecular mimicry between IRRET motive in P. mirabilis urease and human type XI collagen performed on a population in Mexico, revealed that the per- (LRREI sequence) present in hyaline cartilage. The observed centage of infected individuals increases with age. The pres- ence of anti-urease IgG antibodies in serum rises from less similarities concerned a sequence as well as a conformation fragments of both proteins. Simultaneously, the level of anti- than 20% in a group of individuals below 10 years old to bodies against P. mirabilis urease was significantly higher in more than 50% - in a group over 40 years old [93]. Occur- comparison to healthy individuals as well as patients with rence of anti-urease antibodies was correlated with disease ankylosing spondylitis - another autoimmune disease. Ac- severity. In patients with superficial gastritis, a low level of cording to Wilson et al. hypothesis, antibodies arising in IgG, but relatively high of IgA immunoglobulins was ob- reaction against bacterial urease function as autoantibodies served. Strong IgG reaction dominated in quiescent atrophic and recognize also human protein (collagen). This leads to gastritis individuals, whereas in patients with active atrophic primary cytotoxic damage to hyaline cartilage. In the next gastritis, reaction of IgG as well as IgA was very strong [15]. step, in an injury site the presence of cytokines, vascular Nurgalieva et al. observed the presence of IgM antibod- adhesion molecules and hydrolytic enzymes is observed. It ies, putatively recognizing a small subunit (UreA) of H. py- causes inflammation, fibrosis and destruction of joints [101]. lori urease in 94% of H. pylori-infected volunteers. The This hypothesis was confirmed in later studies. larger subunit - UreB seems to be less immunogenic. About Konieczna et al. observed for RA patient’s sera a signifi- 44% of the investigated individuals showed a positive reac- cantly higher level of antibodies recognizing synthetic pep- tion [94]. However, Burnie and Al-Dughaym showed that tide corresponding to flap epitope of P. mirabilis urease. UreB subunit of H. pylori urease has more epitopes recog- Surprisingly, they noted an elevated IgG level against pep- nized by antibodies than UreA. The level of IgG antibodies tides reflecting a sequence of flap regions from other organ- recognizing some of those epitopes was comparable with the isms (bacteria and plant). The detected antibodies also had commercial test [95]. Also in Arabski et al. study, where lower specificity. These antibodies recognized not only one levels of IgG antibodies were detected, the presence of anti- defined antigen, but also antigens with a similar sequence, bodies recognizing H. pylori UreB urease subunit was found which was probably due to instability of the immune system in almost all infected individuals. They were observed even [7]. in 70% of H. pylori negative sera. A much more interesting A role of urease in stimulation of immune response of observation was a correlation between atherosclerosis and patients with immune disease was also revealed for other the presence of anti-urease antibodies. In the investigated gram-negative bacteria. In 1993, it was showed that sera there was a significant relationship between the level of subunit of Y. enterocilitica O:3 urease is arthritogenic for antibodies bound to 8-mer synthetic peptide (which corre- rats [84]. A few years later, a high humoral response in pa- sponds to UreB minimal flap epitope of H. pylori urease) and tients with reactive arthritis triggered by Y. enterocolitica occurrence of atherosclerosis [6], an inflammatory disease O:3 was noted. IgG reacting with 19 kDa urease subunit was leading to an atheromatosus plaque in blood vessels lumen observed in over 90%, and IgA in over 50 % of investigated [96]. Earlier, Oshima et al. suggested that chronic H. pylori sera [102]. infections are connected with inflammatory processes in vessels [97]. Investigations applying synthetic peptide corre- In chronic obstructive pulmonary disease (COPD) caused sponding to 8 amino acid sequence of flap fragment of H. by nonencapsulated H. influenzae, urease is a target of hu- pylori urease revealed a similarity between this peptide and man humoral response. In almost 39% of investigated sera, a human CCRL1 (CC chemokine receptor-like 1) protein, ex- significantly higher level of antibodies reacting with bacte- pressed mainly in the heart. Based on this observation, a hy- rial urease was observed [103]. pothesis to explain a connection of H. pylori urease and athe- Anti-urease antibodies are detected even in case of rosclerosis was formulated. According to it, urease may chronic zoonosis. In patients with diagnosed brucellosis, stimulate immune system reaction during a bacterial infec- antibodies recognizing  Brucella suis urease subunit were tion. Presentation of urease fragments to Th lymphocytes detected [104]. enables synthesis of antibodies. Next, antibodies against flap 798 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (7). Molecular mimicry of flap fragment of H. pylori urease and CCRL1 and possible connection with atherosclerosis progress. In the investigations of antibodies generated as a re- sembled), apart from continuous (sequential, linear), epitopes sponse to infection, chemically defined synthetic peptides [106]. However, several strategies of epitope mapping are have a potent application. They are useful for epitope map- available. The most often used strategy is array-based oligo- ping as well as molecular mimicry studies. peptide scanning. This technique uses a library of oligo- peptide sequences from overlapping and non-overlapping 8. ORGANIC CHEMISTRY TOOLS IN IMMUNE RE- segments of a target protein and tests for their ability to bind SPONSE INVESTIGATIONS the antibody of interest. This method is fast and relatively inexpensive, and specifically suited to profile epitopes for a Organic chemistry enables constructing several new tools large number of candidate antibodies against defined targets for investigations of response of the immune system against [108]. infectious agents. One of these tools is a library of synthetic peptides with a chemically defined sequence. Such libraries So far the most general approach for epitope mapping has are used for detecting antibodies as well as for estimating been developed by Ronald Frank [109]. The applied meth- their variety and specificity [105]. Synthetic peptide libraries odology, known as SPOT synthesis, is a special type of solid also provide epitope mapping of protein antigens, the process phase peptide synthesis proceeding directly on the membrane of locating the epitope on the protein surface or in the protein support, inside relatively small, separated spots regarded as sequence [106]. There are many peptide synthesis methods: separate reaction vessels. The method was initiated as an biological (peptide is expressed on the surface of bacterium uncomplicated manual technique for parallel chemical syn- or phage) or fully synthetic (peptide is synthesized on the thesis of peptide arrays followed by an assay with appropri- abiotic surface like cellulose or polypropylene) [107]. ate interacting molecules performed directly on the mem- brane or in solution (Fig. 8) [105]. Determination of antigenic determinants of protein may be intricate considering the existence of discontinuous (as- Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 799 Fig. (8). Scheme of SPOT synthesis. SPOT method is useful for mapping not only linear, but Synthesis of peptide on the membrane surface allows also discontinuous epitopes as well as for characterizing an- application of simple techniques, like dot-blot, for further tibodies [110, 111, 112]. It was even applied for identifica- investigations of e.g. bounded antibodies. However, the re- tion of peptide mimicking the structure of an epitope (mimo- sults obtained strongly depend on the structure of the linker tope) [110]. A broad variety of other biomolecular binding fragment. The most classical linkers are prepared using 1-3 events or enzymatic modifications can be investigated by residues of  -alanine or glycine to separate peptide assem- using peptide arrays (prepared by the SPOT technique) such bling with biomolecules used in assay out of the membrane as protein-protein interactions [113-118] protein-DNA inter- [109, 128]. In SPOT technology, different linkers are also actions [119] peptide-cell interactions [120-122] or enzyme- applied (like Carboxy-Frank-Linker, p-hydroxy- substrate interactions [123-127]. methylbenzoic acid (HMB) linker, the Rink-amide linker or 800 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (9). Synthesis of peptides with free N-termini anchored by isocyanuric linker. 4-hydroxymethyl-phenoxy acetic acid (HMPA) and 4-(4- A different type of anchoring the peptide chain to cellu- hydroxymethyl-3-methoxyphenoxy)-butyric acid (HMPB) lose matrix was proposed by Kaminski and co-workers [6, linkers) enabling cleavage of peptides from the support [129- 134-136]. 1-Acyl-3,5-dimethyl-1,3, 5-triazin-2,4,6 (1H,3H, 133]. 5H)-trion derivatives serve both as a spacer and a linker. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 801 This isocyanuric linker has been introduced by thermal iso- on the synthesis of the target amino acid sequence as a series merization of 2-acyloxy-4,6-dimethoxy-1,3,5-triazines al- of overlapping peptides on polypropylene pins [30]. ready immobilized on the cellulose support [137]. A syn- Studies on urease epitopes were also performed using thetic procedure leading to peptides anchored to cellulose by other techniques. One of the most interesting new develop- 1-acyl-3,5-dimethyl-1,3,5-triazin-2,4,6(1H,3H,5H)-trion ments in the search for novel antigens applied a computa- (iso-MT) is shown in (Fig. 9). In the first step, chloro- tional method to predict T-cell epitopes using the whole ge- triazine immobilized on cellulose was treated with N- nome sequence information [128]. methylmorpholine yielding N-triazinylammonium chloride, In a more classical approach, antibody binding epitopes which is activating the carboxylic function of Fmoc- in H. pylori urease were determined with monoclonal anti- protected amino acid yielding superactive triazine type ester [135, 136], which, in refluxing toluene, rearranges to a stable bodies produced in Balb/c mice. After digestion of bacterial isocyanuric derivative. urease with trypsin followed by separation of peptides with affinity chromatography, monoclonal antibodies were ap- During peptide synthesis, anchoring method is essential plied to identify epitopes. Next, amino acid sequences of for further reaction of peptide and antibody (Table 4). isolated peptides were determined by mass spectroscopic For isocyanuric linker, interactions with antibodies were analysis. Fujii et al. showed that two such peptides, with the found more selective [138]. This linker was applied during sequence SVELIDIGGNRRIFGFNALVDR and IFGFNAL synthesis of peptides corresponding to a flap region of dif- VDR, were recognized by two monoclonal antibodies ferent ureases (bacterial and plant) based on the flap region (MAb): HpU-2 and HpU-18 respectively. Based on the data from H. pylori urease. Those peptides were useful in investi- of competitive binding determined by using surface plasmon gating human sera. It was possible to differentiate sera of resonance and analysis of the epitope for HpU-2 and HpU- patients with autoimmune diseases like rheumatoid arthritis 18, it has been found that both MAb recognize almost the or atherosclerosis from sera of healthy donors [6, 7]. On the same position on the UreA subunit of H. pylori urease. An other hand, a library of 361 peptides, where each peptide had unanticipated result was suppressing of urease activity via an a sequence with one substituted amino acids in a defined allosteric effect, which might cause a distortion of the con- position, was applied to determine amino acids which are formation of the enzyme. On the other hand, the second essential for antibody binding [139]. mechanism observed in the case of several MAb studied For epitope mapping of UreA and UreB subunits of H. (which in fact possessed a weaker inhibitory effect, such as pylori urease, Geysen’s method was adopted, which is based HpU-17 and -20) is assumed that MAb B binds to the vicin- Table 4. Interaction of Antibodies with H. pylori Urease Epitopes with Free N-Termini Anchored on Cellulose. Epitope Peptide sequence Reaction Specificity cellulose H C N (Aaa) -NH n 2 HN-CHHLDKSIKEDVQFADSRI-COO-cellulose - UB-33 H N-CHHLDKSIKEDVQFADSRI--Ala-COO-cellulose - 0% H N-CHHLDKSIKEDVQFADSRI- -Ala- -Ala--Ala-COO-cellulose - cellulose N N Me (Aaa) -NH n 2 H N-CHHLDKSIKEDVQFADSRI--Ala-iso-MT-cellulose +, m UB-33 H2N-CHHLDKSIKEDVQFADSRI-iso-MT-cellulose +, s 100% H N-SIKEDVQF--Ala-iso-MT-cellulose +, s F-8 HN-SIKEDVQF-iso-MT-cellulose +, m a) - (no reaction); + (reaction); s (strong); m (medium). 802 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. ity of the active site, resulting in the reduction of the urease Urease, although investigated for a long time, still seems to be an unexplored enzyme. activity [140]. Molecular biology methods are also used in H. pylori CONFLICT OF INTEREST urease epitope determination. Nineteen truncated fragments of gene coding UreB subunit were amplified and cloned into The author(s) confirm that this article content has no con- the prokaryotic expression vector pET-28a (+) or pGEX-4T- flicts of interest. 2. After verification, the constructs obtained were trans- ACKNOWLEDGEMENTS formed into Escherichia coli which expressed recombinant proteins. Using three MAbs against UreB of H. pylori This work was supported by grant N N304 044639 from (A1H10, A3C10, and B3D9) three linear B-cell epitopes, Ministry of Sciences and High Education (Poland). probably useful as the targets for development of epitope- based vaccines against H. pylori, were identified. These epi- REFERENCES topes were localized in the aa regions: 158-172 [1] Mobley, H.L.T.; Island, M.D.; Hausinger, R.P. Molecular biology (GGGTGPADGTNATTI), 181-195 (WMLRAAEEYS of microbial ureases. Microbiol. Rev., 1995, 59(3), 451-480. MNLGF), and 349-363 (TLHDMGIFSITSSDS) of UreB [2] Newsholme, E.; Leech, A. Functional Biochemistry in Health and nd Disease, 2 ed.; John Wiley & Sons, Chichester, 2011. [141]. [3] Sirko, A.; Brodzik, R. Plant ureases: Roles and regulation. Acta Urease is highly expressed by all strains of H. pylori and is Biochim. Pol., 2000, 47(4), 1189-1195. immunogenic. Additionally this enzyme could stimulate [4] Dunn, B.E.; Phadnis S.H. Structure, Function and Localization of Helicobacter pylori Urease. Yale J. Biol. Med., 1998, 71(2), 63-73. generation of antibodies able to inhibit its activity. For this [5] Follmer, C. Ureases as a target for the treatment of gastric and reason, it seems to be a promising vaccine target. However, urinary infections. J. Clin. Pathol., 2010, 63(5), 424-430. vaccination of urease may not give a sufficient protective [6] Arabski, M.; Konieczna, I.; So owiej, D.; Rogo , A.; Kolesi ska, effect. A combination of urease with other antigens may B.; Kami ski, Z.; Kaca, W. Are anti-H. pylori urease antibodies in- yield better results. It seems that a fusion of UreB urease volved in atherosclerotic disease? Clin. Biochem., 2010, 43(1-2), subunit with truncated HpaA surface protein may give a bet- 115-123. [7] Konieczna, I.; Kwinkowski, M.; Kolesi ska, B.; Kami ski, Z.; ter protection than either protein alone [142]. Fr czyk, J.; arnowiec, P.; Kaca, W. Detection of Antibodies There were also attempts to design a vaccine using mu- against Synthetic Peptides Mimicking Ureases Fragments in Sera cosal adjuvant cholera toxin B subunit (CTB) and an epitope of Rheumatoid Arthritis Patients. Prot. Pept. Lett., 2012, 19(11), 1149-1154. (UreA 183-203) of H. pylori urease. Both peptides were [8] Mobley, H.L.T.; Hausinger, R.P. Microbial ureases: significance, bound with the linker (DPRVPSS) to avoid the formation of regulation, and molecular characterization. Microbiol. Rev., 1989, new epitopes. The CTB-UreA epitope vaccine had good im- 53(1), 85-108. munogenicity and immunoreactivity and induced specific [9] Suzuki, K.; Benno, Y.; Mitsuoka, T.; Takebe, S.; Kobashi, K.; neutralizing antibodies which showed an effectively inhibi- Hase, J. Urease-Producing Species of Intestinal Anaerobes and tory effect on H. pylori urease enzymatic activity [143]. Their Activities. Appl. Environ. Microbiol., 1979, 37(3), 379-382. [10] Murchan, S.; Aucken, H.M.; O’Neill, G.L.; Ganner, M.; Cookson, Today, experiments to identify and characterize linear B.D. Emergence, Spread, and Characterization of Phage Variants antibody epitopes using peptide scans, amino acids scans, of Epidemic Methicillin-Resistant Staphylococcus aureus 16 in substitutional analyses, truncation libraries, deletion librar- England and Wales. J. Clin. Microbiol., 2004 42(11), 5154-5160. [11] Jin, M.; Rosario, W.; Watler, E.; Calhoun, D.H. Development of a ies, cyclization scans, all types of combinatorial libraries and large-scale HPLC-based purification for the urease from Staphylo- randomly generated libraries of single peptides are standard coccus leei and determination of subunit structure. Protein Expr. techniques widely applied even in non-specialized laborato- Purif. 2004, 34(1), 111-117. ries [135]. [12] Clemens, D.L.; Lee, B-Y.; Horwitz, M.A. Purification, Characteri- zation, and Genetic Analysis of Mycobacterium tuberculosis CONCLUSIONS Urease, a Potentially Critical Determinant of Host-Pathogen Inter- action. J. Bacteriol., 1995, 177(19), 5644-5652. Urease is an enzyme studied for a long time. Its structure, [13] Dupuy, B.; Daube, G.; Popoff, M.R.; Cole, S.T. Clostridium per- fringens Urease Genes Are Plasmid Borne. Infect. Immun., 1997, synthesis and biochemical activity are known. There are also 65(6), 2313-2320. many studies concerning urease toxic effect on human tis- [14] Lam, S.; Yeo, M. Urease-Positive Vibrio parahaemolyticus Strain. sues. However, its role in long-lasting autoimmune diseases J. Clinical Microbiol., 1980, 12(1), 57-59. is still controversial. Nevertheless, the presence of molecular [15] Futagami, S.; Takahashi, H.; Norose, Y.; Kobayashi, M. Systemic mimicry between bacterial ureases and human proteins has and local immune responses against Helicobacter pylori urease in patients with chronic gastritis: distinct IgA and IgG productive been suggested [7, 101]. Proteins containing motives, similar sites. Gut, 1998, 43(2), 168-175. to infectious agents, may function as autoantigens. In de- [16] Nakano, M.; Iida, T.; Ohnishi, M.; Kurokawa, K.; Takahashi, A.; scribed autoantigenes, some similarities to ureases may be Tsukamoto, T.; Yasunaga, T.; Hayashi, T.; Honda, T. Association found [90, 101]. It was proved that this enzyme stimulates of the Urease Gene with Enterohemorrhagic Escherichia coli antibodies synthesis [89], but determination of epitopes in Strains Irrespective of Their Serogroups. J. Clin. Microbiol., 2001, urease protein may be difficult and non-conclusive. There- 39(12), 4541-4543. [17] Orth, D.; Grif, K.; Dierich, M.P.; Würzner, R. Prevalence, structure fore investigations applying synthetic peptides could be very and expression of urease genes in Shiga toxin-producing Es- helpful in mapping epitopes both in infectious agents pro- cherichia coli from humans and the environment. Int. J. Hyg. Envi- teins as well as in determining amino acids located in epi- ron.-Health, 2006, 209(6), 513-520. topes which are essential for human humoral response [139]. [18] Tange, Y.; Niwa, O. Identification of the ure1+ gene encoding urease in fission yeast. Curr. Genet., 1997, 32(3), 244-246. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 803 [19] Yu, J.J.; Smithson, S.L.; Thomas, P.W.; Kirkland, T.N.; Cole, G.T. that are competent for in vivo enzyme activation. J. Bacteriol., Isolation and characterization of the urease gene (URE) from the 1995, 177(8), 1947-1951. pathogenic fungus Coccidioides immitis. Gene, 1997, 198(1-2), [40] Sriwanthana, B.; Island, M.D.; Maneval, D.; Mobley, H.L.T. Single 387-391. step purification of Proteus mirabilis urease accessory protein [20] Zonia, L.E.; Stebbins, N.E.; Polacco, J.C. Essential role of urease UreE, a protein with a naturally occurring histidine tail, by nickel in germination of nitrogen-limited Arabidopsis thaliana seeds. chelate affinity chromatography. J. Bacteriol., 1994, 176(22), Plant Physiol., 1995, 107(4), 1097-1103. 6836-6841. [21] Pedrozo, H.A.; Schwartz, Z.; Luther, M.; Dean, D.D.; Boyan, B.D.; [41] Sriwanthana, B.; Island, M.D.; Mobley, H.L.T. Sequence of the Wiederhold, M.L. A mechanism of adaptation to hypergravity in Proteus mirabilis urease accessory gene ureG. Gene, 1993, 129(1), the statocyst of Aplysia californica. Hear Res., 1996, 102(1-2), 51- 103-106. 62. [42] Voland, P.; Weeks, D.L.; Marcus, E.A.; Prinz, C.; Sachs, G.; Scott, [22] Hedelin, H. Uropathogens and urinary tract concretion formation D. Interactions among the seven Helicobacter pylori proteins en- and catheter encrustations. Int. J. Antimicrob. Agents, 2002, 19(6), coded by the urease gene cluster. Am. J. Physiol. Gastrointest. 484-487. Liver Physiol., 2003, 284(1), 96-106. [23] Carter, E.L.; Proshlyakov, D.A.; Hausinger, R.P. Apoprotein isola- [43] Park, I.-S.; Hausinger, R.P. Requirement of CO2 for in vitro assem- tion and activation, and vibrational structure of the Helicobacter bly of the urease nickel metallocenter. Science, 1995, 267(5201), mustelae iron urease. J. Inorg. Biochem., 2011, 111, 195-202. 1156-1158. [24] Rózalski, A.; Kwil, I.; Torzewska, A.; Baranowska, M.; Staczek, P. [44] Musiani, F.; Zambelli, B.; Stola, M.; Ciurli, S. Nickel trafficking: Proteus bacilli: features and virulence factors. Postepy Hig. Med. insights into the fold and function of UreE, a urease metallochaper- Dosw., 2007, 61, 204-219. one. J. Inorg. Biochem., 2004, 98(5), 803-813. [25] Lv, J.; Jiang, Y.; Yu, Q.; Lu, S. Structural and functional role of [45] Palinska, K.A.; Jahns, T.; Rippka, R.; Tandeau de Marsac, N. Pro- nickel ions in urease by molecular dynamics simulation. J Biol chlorococcus marinus strain PCC 9511, a picoplanktonic cyano- Inorg Chem., 2011, 16(1), 125-135. bacterium, synthesizes the smallest urease. Microbiology, 2000, [26] Mörsdorf, G.; Kaltwasser, H. Cloning of the genes encoding urease 146, 3099-3107. from Proteus vulgaris and sequencing of the structural genes. [46] Ha, N.-C.; Oh, S-T.; Sung, J.Y.; Cha, K.A.; Lee, M.H.; Oh, B.-H. FEMS Microbiol. Lett., 1990, 54(1-3), 67-73. Supramolecular assembly and acid resistance of Helicobacter py- [27] Walz, S.E.; Wray, S.K.; Hull, S.I.; Hull, R.E. Multiple proteins lori urease. Nat. Struct. Biol., 2001, 8, 505-509. encoded within the urease gene complex of Proteus mirabilis. J. [47] McMillan, D.J.; Mau, M.; Walker, M.J. Characterisation of the Bacteriol., 1988, 170(3), 1027-1033. urease gene cluster in Bordetella bronchiseptica. Gene, 1998, [28] Jones, B.D.; Mobley, H.L.T. Proteus mirabilis urease: nucleotide 208(2), 243-251. sequence determination and comparison with jack bean urease. J. [48] Bosse, J.T.; MacInnes, J.I. Genetic and Biochemical Analyses of Bacteriol., 1989, 171(12), 6414-6422. Actinobacillus pleuropneumoniae Urease. Infect. Immun., 1997, [29] Schoep, T.D.; Fulurija, A.; Good, F.; Lu, W.; Himbeck, RP, 65(11), 4389-4394. Schwan C.; Choi S. S.; Berg D.E.; Mittl P.R.E.; Benghezal M.; [49] Monteiro, L.; de Mascarel, A.; Sarrasqueta, A.M.; Bergey, B.; Marshall B.J. Surface Properties of Helicobacter pylori Urease Barberis, C.; Talby, P.; Roux, D.; Shouler, L.; Goldfain, D.; Complex Are Essential for Persistence. PLoS ONE, 2010, 5(11), Lamouliatte, H.; Megraud, F. Diagnosis of Helicobacter pylori in- e15042. doi:10.1371/journal.pone.0015042 fection: noninvasive methods compared to invasive methods and [30] Clayton, C.L.; Pallen, M.J.; Kleanthous, H.; Wren, B.W.; Ta- evaluation of two new tests. Am. J. Gastroenterol., 2001, 96(2), baqchali, S. Nucleotide sequence of two genes from Helicobacter 353-358. pylori encoding for urease subunits. Nucleic Acids Res., 1990, [50] Falsafi, T.; Favaedi, R.; Mahjoub, F.; Najafi M. Application of 18(2), 362. Stool-PCR test for diagnosis of Helicobacter pylori infection in [31] Ferrero, R.L.; Labigne, A. Cloning, expression and sequencing of children World J. Gastroenterol., 2009, 15(4), 484-488. Helicobacter felis urease genes. Mol. Microbiol., 1993, 9(2), 323- [51] Ragsdale, S.W. Nickel-based Enzyme Systems. J. Biol. Chem., 333. 2009, 284(28), 18571-18575. [32] Labigne, A.; Cussac, V.; Courcoux, P. Shuttle cloning and nucleo- [52] Konieczna, I. Characteristic of the bacterial ureases molecular tide sequences of Helicobacter pylori genes responsible for urease variety and estimation of reactivity of the human anti-urease anti- activity. J. Bacteriol., 1991, 173(6), 1920-1931. bodies. PhD Thesis, The Jan Kochanowski University of Humani- [33] Solnick, J.V.; O’Rourke, J.; Lee, A.; Tompkins, L.S. Molecular ties and Sciences: Kielce, April, 2010. analysis of urease genes from a newly identified uncultured species [53] Hamilton-Miller, J.M.T.; Gargan, R.A. Rapid screening for urease of Helicobacter. Infect. Immun., 1994, 62(5), 1631-1638. inhibitors. Invest. Urol., 1979, 16(5), 327-328. [34] Lee, S.G.; Calhoun, D.H. Urease from a Potentially Pathogenic [54] Christensen, W.B. Urea decomposition as a means of differentiat- Coccoid Isolate: Purification, Characterization, and Comparison to ing Proteus and paracolon cultures from each other and from Sal- Other Microbial Ureases. Infect. Immun., 1997, 65(10), 3991-3996. monella and Shigella types. J. Bacteriol., 1946, 52(4), 461-466. [35] Lee, M.H.; Mulrooney, S.B.; Renner, M.J.; Markowicz, Y.; Haus- [55] Hussain Qadri, S.M.; Zubairi, S.; Hawley, H.P.; Mazlaghani, H.H.; inger, R.P. Klebsiella aerogenes urease gene cluster: sequence of Ramirez, E.G. Rapid test for determination of urea hydrolysis. An- ureD and demonstration that four accessory genes (ureD, ureE, tonie van Leeuwenhoek, 1984, 50(4), 417-423. ureF, ureG) are involved in nickel metallocenter biosynthesis. J. [56] Bell, G.D.; Weil, J.; Harrison, G.; Morden, A.; Jones, P.H.; Gant, Bacteriol., 1992, 174(13), 4324-4330. P.W.; Trowell, J.E.; Yoong, A.K.; Daneshmend, T.K.; Logan, [36] Lee, M.H.; Pankratz, H.S.; Wang, S.; Scott, R.A.; Finnegan, M.G.; R.F.A. 14C-urea breath analysis, a non-invasive test for Campy- Johnson, M.K.; Ippolito, J.A.; Christianson, D.W.; Hausinger, R.P. lobacter pylori in the stomach. Lancet, 1987, 329(8546), 1367- Purification and characterization of Klebsiella aerogenes UreE pro- 1368. tein: a nickel-binding protein that functions in urease metallocenter [57] Vakil, N.; Rhew, D.; Soll, A.; Ofman, J.J. The costeffectiveness of assembly. Protein Sci., 1993, 2(6), 1042-1052. diagnostic testing strategies for Helicobacter pylori. Am. J. Gastro- [37] Mulrooney, S.B.; Hausinger, R.P. Sequence of the Klebsiella aero- enterol., 2000, 95(7), 1691-1698. genes urease genes and evidence for accessory proteins facilitating [58] Mansour-Ghanaei, F.; Sanaei, O.; Joukar, F. Clinical Validation of nickel incorporation. J. Bacteriol., 1990, 172(10), 5837-5843. an Office-Based 14C-UBT (Heliprobe) for H. pylori Diagnosis in [38] Park, I.-S.; Carr, M.B.; Hausinger, R.P. In vitro activation of urease Iranian Dyspeptic Patients. Gastroenterol. Res. Pract., 2011, 2011, apoprotein and role of UreD as a chaperone required for nickel 930941. metallocenter assembly. Proc. Natl. Acad. Sci. USA, 1994, 91(8), [59] Foroutan, M.; Loloei, B.; Irvani, S.; Azargashb, E. Accuracy of 3233-3237. rapid urease test in diagnosing Helicobacter pylori infection in pa- [39] Park, I.-S.; Hausinger, R.P. Evidence for the presence of urease tients using NSAIDs. Saudi. J. Gastroenterol., 2010, 16(2), 110- apoprotein complexes containing UreD, UreF, and UreG in cells 112. 804 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. [60] Krogfelt, K.A.; Lehours, P.; Megraud, F. Diagnosis of Helicobac- [83] Coker, C.; Poore, C.A.; Li, X.; Mobley, H.L.T. Pathogenesis of ter pylori Infection. Helicobacter, 2005, 10(Suppl 1), 5-13. Proteus mirabilis urinary tract infection. Microbes Infect., 2000, [61] Artiko, V.M.; Obradovi , V.B.; Petrovi , N.S.; Davidovi , B.M.; 2(12), 1497-1505. Gruji -Adanja, G.S.; Nasti -Miri , D.R.; Milosavljevi . T.N. 14C- [84] Probst, P.; Hermann, E.; Meyer Zum Buschenfelde, K.-H.; urea breath test in the detection of Helicobacter pylori infection. Fleischer, B. Identification of the Yersinia enterocolitica Urease Nuc. Med. Rev., 2001, 4(2), 101-103. Subunit as a Target Antigen for Human Synovial T Lymphocytes [62] Jassal, M.S.; Nedeltchev, G.G.; Lee, J-H.; Choi, S.W.; Atudorei, in Reactive Arthritis. Infect. Immun., 1993, 61(10), 4507-4509. V.; Sharp, Z.D.; Deretic, V.; Timmins, G.S.; Bishai, WR. [C]- [85] Wroblewski, L.E.; Shen, L.; Ogden, S.; Romero-Gallo, J.; Lapierre, Urea Breath Test as a Novel Point-of-Care Biomarker for Tubercu- L.A.; Israel, D.A.; Turner, J.R.; Peek, R.M.Jr. Helicobacter pylori losis Treatment and Diagnosis. PLoS ONE, 2010, 5(8), e12451. dysregulation of gastric epithelial tight junctions by urease- [63] Ruiz-Herrera, J.; Gonzalez, J. A continuous method for the meas- mediated myosin II activation. Gastroenterology, 2009, 136(1), urement of urease activity. Anal. Biochem., 1969, 31, 366-374. 236-246. [64] Blattler, D.P.; Contaxis, C.C.; Reithel, F.J. Dissociation of urease [86] Wassermann, G.E.; Olivera-Severo, D.; Uberti, A.F.; Carlini, C.R. by glycol and glycerol. Nature, 1967, 216, 274-275. Helicobacter pylori urease activates blood platelets through a [65] Shaik-M, M.B.; Guy, A.L.; Pancholy, S.K. An improved method lipoxygenase-mediated pathway. J. Cell. Mol. Med., 2010, 14(7), for the detection and preservation of urease activity in polyacryla- 2025-2034. mide gel. Anal. Biochem., 1980, 103, 140-143. [87] Johnson, D.E.; Russell, R.G.; Lockatell, C.V.; Zulty, J.C.; Warren, [66] Weatherburn, M.W. Phenol-hypochlorite reaction for determination J.W.; Mobley, H.L.T. Contribution of Proteus mirabilis Urease to of ammonia. Anal. Chem., 1967, 39, 971-974. Persistence, Urolithiasis, and Acute Pyelonephritis in a Mouse [67] Krajewska, B.; Chudy, M.; Drozdek, M.; Brzozka, Z. Potentiomet- Model of Ascending Urinary Tract Infection Infect. Immun., 1993, ric Study of Urease Kinetics over pH 5.36 -8.21. Electroanalysis, 61(7), 2748-2754. 2003, 15(5-6), 460-466. [88] Burne R.A.; Chen, Yi-Y.M. Bacterial ureases in infectious dis- [68] Juszkiewicz, A.; Kot, M.; Zaborska, W. Calorimetric study of eases. Microbes Infect., 2000, 2(5), 533-542. inhibition of urease by 2-mercaptoethanol. Procedures based upon [89] Hirota, K.; Nagata, K.; Norose, Y.; Futagami, S.; Nakagawa, Y.; integrated rate equations. Thermochim. Acta, 1998, 320(1-2), 45- Senpuku, H.; Kobayashi, M.; Takahashi, H. Identification of an an- 52. tigenic epitope in Helicobacter pylori urease that induces neutraliz- [69] Lin, Y-L.; Chen, Ch-T.; Lin, S-C.; Lee, C.; Kuo, H-S.; Shih, C-M.; ing antibody production. Infect. Immun., 2001, 69(11), 6597-6603. Hsu, Y-H.; Chin, Y-P.; Chan, E-C. A simple method to determine [90] Backes, C.; Ludwig, N.; Leidinger, P.; Harz, C.; Hoffmann, J.; urea concentration using intact Helicobacter pylori and Bromo Keller, A.; Meese, E.; Lenhof, H-P. Immunogenicity of autoanti- Cresol Purple as a pH indicator. Biotechnol. Lett., 2000, 22(13), gens. BMC Genom., 2011, 12, 340. 1077-1079. [91] Thomas, J.E.; Whatmore, A.M.; Barer, M.R.; Eastham, E.J.; Ke- [70] Kaltwasser, H.; Schlegel, H.G.. NADH-dependent coupled assay hoe, M.A. Serodiagnosis of Helicobacter pylori Infection in Child- for urease and other ammonia-producing systems. Anal. Biochem., hood. J. Clin. Microbiol., 1990, 28(12), 2641-2646. 1966, 16(1), 132-138. [92] Dunn, B.E.; Cohen, H.; Blaser, M.J. Helicobacter pylori. Clin. [71] Gambhir, A.; Gerard, M.; Mulchandani, A.K.; Malhotra, B.D. Microbiol. Rev., 1997, 10(4), 720-741. Coimmobilization of Urease and Glutamate Dehydrogenase in [93] Leal-Herrera, Y.; Torres, J.; Perez-Perez, G.; Gomez, A.; Monath, Electrochemically Prepared Polypyrrole-Polyvinyl Sulfonate Films. T.; Tapia-Conyer, R.; Muñoz, O. Serologic IgG Response To Appl. Biochem. Biotechnol., 2001, 96(1-3), 249-257. Urease In Helicobacter pylori-Infected Persons From Mexico Am. [72] Stutts, P.; Fridovich, I. A continual spectrophotometric determina- J. Trop. Med. Hyg., 1999, 60(4), 587-592. tion of ammonia-producing systems. Anal. Biochem., 1964. 8, 70- [94] Nurgalieva, Z.Z.; Conner, M.E.; Opekun, A.R.; Zheng, C.Q.; 74. Elliott, S.N.; Ernst, P.B.; Osato, M.; Estes, M.K.; Graham, D.Y. B- [73] Katz, S.A. Direct potentiometric determination of urease activity. Cell and T-Cell Immune Responses to Experimental Helicobacter Anal. Chem., 1964, 36(13), 2500-2501. pylori Infection in Humans. Infect. Immun., 2005, 73(5), 2999- [74] Montalvo, J.C., Jr. An improved urease electrode. Anal. Biochem., 3006. 1970, 36(2), 357-363. [95] Burnie, J.P.; Al-Dughaym, A. The application of epitope mapping [75] McDonald, J.A.; Speeg, K.V.; Campbell, J.W. Urease: a sensitive in the development of a new serological test for Helicobacter pylori and specific radiometric assay. Enzymologia, 1972, 42, 1-9. infection. J. Immunol. Methods, 1996, 194(1), 85-94. [76] Wrong, O.M.; Vince, A.J.; Waterlow, J.C. The contribution of [96] Milioti, N.; Bermudez-Fajardo, A.; Penichet, M.L.; Ernesto, O.-O. endogenous urea to face alammonia in man, determined by 15N- Antigen-Induced Immunomodulation in the Pathogenesis of Athe- labeling of plasma urea. Clin. Sci., 1985, 68(2), 193-199. rosclerosis. Clin. Dev. Immunol., 2008, 2008, 723539. doi: [77] Solomon, C.M.; Alexander, J.A.; Glibert P.M. Measuring urease 10.1155/2008/723539. activity in aquatic environmental samples. Limnol. Oceanogr. [97] Oshima, T.; Ozono, R.; Yano, Y.; Oishi, Y.; Teragawa, H.; Higa- Methods, 2007, 5, 280-288. shi, Y.; Yoshizumi, M.; Kambe, M. Association of Helicobacter [78] Karmali, K.; Karmali, A.; Teixeira, A.; Curto, M.J. The use of pylori Infection With Systemic Inflammation and Endothelial Dys- Fourier transform infrared spectroscopy to assay for urease from function in Healthy Male Subjects. J. Am. Coll. Cardiol., 2005, Pseudomonas aeruginosa and Canavalia ensiformis. Anal. Bio- 45(8), 1219-1222. chem., 2004, 331(1), 115-121. [98] Aletaha, D.; Neogi, T.; Silman, A.J.; Funovits, J.; Felson, D.T.; [79] Kumar, S.; Barth, A. Following Enzyme Activity with Infrared Bingham, C.O.; Birnbaum, N.S.; Burmester, G.R.; Bykerk, V.P.; Spectroscopy. Sensors, 2010, 10, 2626-2637. Cohen, M.D.; Combe, B.; Costenbader, K.H.; Dougados, M.; Em- [80] Olivera-Severo, D.; Wassermann1, G.E.; Carlini, C.R. Ureases ery, P.; Ferraccioli, G.; Hazes. J.M.W.; Hobbs, K.; Huizinga, display biological effects independent of enzymatic activity. Is T.W.J.; Kavanaugh, A.; Kay, J.; Kvien, T.K.; Laing, T.; Mease, P.; there a connection to diseases caused by urease-producing bacteria? Menard, H.A.; Moreland, L.W.; Naden, R.L.; Pincus, T.; Smolen, Braz. J. Med. Biol. Res., 2006, 39(7), 851-861. J.S.; Stanislawska-Biernat, E.; Symmons, D.; Tak, P.P.; Upchurch, [81] Lin, W.; Mathys, V.; Ang, E.L.; Koh, V.H.; Martínez Gómez, J.M.; K.S.; Vencovsk , J.; Wolfe, F.; Hawker, G. 2010 Rheumatoid Ar- Ang, M.L.; Zainul Rahim, S.Z.; Tan, M.P.; Pethe, K.; Alonso, S. thritis Classification Criteria. Arthritis & Rheumatism, 2010, 62(9), Urease activity represents an alternative pathway for Mycobacte- 2569-2581. rium tuberculosis nitrogen metabolism. Infect. Immun., 2012, [99] Rashid, T.; Ebringer, A. Rheumatoid arthritis is linked to Pro- 80(8), 2771-2779. teus—the evidence. Clin. Rheumatol., 2007, 26(7), 1036-1043. [82] Steyert, S.R.; Kaper, J.B. Contribution of Urease to Colonization [100] Tobón, G.J.; Youinou, P.; Saraux, A. The environment, geo- by Shiga Toxin-Producing Escherichia coli. Infect. Immun., 2012, epidemiology, and autoimmune disease: Rheumatoid arthritis. J. 80(8):2589-2600. Autoimmun. 2010, 35(1), 10-14. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 805 [101] Wilson, C.; Tiwana, H.; Ebringer, A. Molecular mimicry between brane-bound peptide repertoires. J. Biol. Chem., 1999, 274(8), HLA-DR alleles associated with rheumatoid arthritis and Proteus 5213-5221. mirabilis as the aetiological basis for autoimmunity. Microbes In- [120] Kato, R.; Kaga, C.; Kanie, K.; Kunimatsu, M.; Okochi, M.; Honda, fect., 2000, 2(12), 1489-1496. H. Peptide Array-Based Peptide-Cell Interaction Analysis. Mini- [102] Appel, H.; Mertz, A.; Distler, A.; Sieper, J.; Braun, J. The 19 kDa Rev Org. Chem., 2011, 8(2), 171-177. protein of Yersinia enterocolitica O:3 is recognized on the cellular [121] Kato, R.; Kaga, C.; Kunimatsu, M.; Kobayashi, T.; Honda, H. and humoral level by patients with Yersinia induced reactive arthri- Peptide arraybased interaction assay of solid-bound peptides and tis. J. Rheumatol., 1999, 26(9),1964-1971. anchorage-dependant cells and its effectiveness in cell-adhesive [103] Murphy, T.F.; Brauer, A.L. Expression of urease by Haemophilus peptide design. J. Biosci. Bioeng., 2006, 101(6), 485-495. influenzae during human respiratory tract infection and role in sur- [122] Falsey, J.R.; Renil, M.; Park, S.; Li, S.; Lam, K.S. Peptide and vival in an acid environment. BMC Microbiol., 2011, 11(1), 183. small molecule microarray for high throughput cell adhesion and [104] Contreras-Rodriguez, A.; Quiroz-Limon, Jose.; Martins, A.M.; functional assays. Bioconjug. Chem., 2001, 12(3), 346-353. Peralta, H.; Avila-Calderon, E.; Sriranganathan N.; Boyle, S.M.; [123] Thiele, A.; Pösel, S.; Spinka, M.; Zerweck, J.; Reimer, U.; Reineke, Lopez-Merino, A. Enzymatic, immunological and phylogenetic U.; Schutkowski, M. Profiling of Enzymatic Activities Using Pep- characterization of Brucella suis urease. BMC Microbiol., 2008, 8, tide Arrays. Mini-Reviews in Organic Chemistry, 2011, 8(2), 147- 121. 156. [105] Shin, D.-S.; Kim, D.-H.; Chung, W.-J.; Lee, Y.-S. Combinatorial [124] Leung, G.C.; Murphy, J.M.; Briant, D.; Sicheri, F. Characterization Solid Phase Peptide Synthesis and Bioassays. J. Biochem. Mol. of kinase target phosphorylation consensus motifs using peptide Biol., 2005, 38(5), 517-525. SPOT arrays. Methods Mol. Biol., 2009, 570, 187-195. [106] Benjamin, D.C.; Berzofsky, J.A.; East, I.J.; Gurd, F.R.; Hannum, [125] Houseman, B.T.; Huh, J.H.; Kron, S.J.; Mrksich, M. Peptide chips C.; Leach, S.J.; Margoliash, E.; Michael, J.G.; Miller, A.; Prager, for the quantitative evaluation of protein kinase activity. Nat. Bio- E.M.; Reichlin, M.; Sercarz, E.E.; Smith-Gill, S.J.; Todd, P.E.; technol., 2002, 20(3), 270-274. Wilson, A.C. The antigenic structure of proteins: a reappraisal. [126] Tegge, W.J.; Frank, R. Analysis of protein kinase substrate speci- Annu. Rev. Immunol., 1984, 2, 67-101. ficity by the use of peptide libraries on cellulose paper (SPOT- [107] Liu, R.; Enstrom, A.M.; Lam, K.S. Combinatorial peptide library method). Methods Mol. Biol., 1998, 87, 99-106. methods for immunobiology research. Exp. Hematol., 2003, 31(1), [127] Espanel, X.; Huguenin-Reggiani, M.; van Huijsduijnen, R.H. The 11-30. SPOT technique as a tool for studying protein tyrosine phosphatase [108] O’Brien-Simpson, N.M.; Pathirana, R.D.; Paolini, R.A.; Chen, Y- substrate specificities. Protein Sci., 2002, 11(10), 2326-2334. Y.; Veith, P.D.; Tam, V.; Ally, N.; Pike, R.N.; Reynolds, E.C. An [128] Moss, S.F.; Moise, L.; Lee, D.S.; Kim, W.; Zhang, S.; Lee, J; Immune Response Directed to Proteinase and Adhesin Functional Rogers, A.B.; Martin, W.; De Groot A.S. HelicoVax: epitope-based Epitopes Protects against Porphyromonas gingivalis-Induced therapeutic Helicobacter pylori vaccination in a mouse model. Periodontal Bone Loss. J. Immunol., 2005, 175(6), 3980-3989. Vaccine, 2011, 29(11), 2085-2091. [109] Frank, R. Spot-synthesis: an easy technique for the positionally [129] Hoffmann, S.; Frank, R. A new safety-catch peptide-resin linkage addressable, parallel chemical synthesis on a membrane support. for the direct release of peptides into aqueous buffers. Tetrahedron Tetrahedron, 1992, 48(42), 9217-9232. Lett., 1994, 35, 7763-7766;. [110] Mahler, M.; Fritzler, M.J. Epitope specificity and significance in [130] Panke, G.; Frank, R. Improved Preparation of a Safety-Catch systemic autoimmune diseases. Ann. N.Y. Acad. Sci., 2010, 1183, Linker for the Solid Phase Synthesis of Peptide Acids Finally Re- 267-287. leased into Aqueous Buffers. Tetrahedron Lett., 1998, 39(1), 17-18. [111] Schwemmle, M.; Billich, C. The use of peptide arrays for the char- [131] Scharn, D.; Wenschuh, H.; Reineke, U.; Schneider-Mergener, J.; acterization of monospecific antibody repertoires from polyclonal Germeroth, L. Spatially addressed synthesis of amino- and amino- sera of psychiatric patients suspected of infection by Borna disease oxy-substituted 1, 3,5-triazine arrays on polymeric membranes. J. virus. Mol. Divers., 2004, 8(3), 247-250. Comb. Chem., 2000, 2(4), 361-369. [112] Hilpert, K.; Hansen, G.; Wessner, H.; Küttner, G.; Welfle, K.; [132] Rau, H.K.; DeJonge, N.; Haehnel, W. Combinatorial Synthesis of Seifert, M.; Höhne, W. Anti-c-myc antibody 9E10: epitope key po- Four-Helix Bundle Hemoproteins for Tuning of Cofactor Proper- sitions and variability characterized using peptide spot synthesis on ties. Angew. Chem. Int. Ed. Engl., 2000, 39(1), 250-253. cellulose. Protein Eng., 2001, 14(10), 803-806. [133] Ay, B.; Landgraf, K.; Streitz, M.; Fuhrmann, S.; Volkmer, R.; [113] Tong, J.; Elowe, S.; Nash, P.; Pawson, T. Manipulation of EphB2 Boisguerin, P. Using hydroxymethylphenoxy derivates with the regulatory motifs and SH2 binding sites switches MAPK signaling SPOT technology to generate peptides with authentic C-termini. and biological activity. J. Biol. Chem., 2003, 278(8), 6111-6119. Bioorg. Med. Chem. Lett., 2008, 18(14), 4038-4043. [114] Yaffe, M.B.; Rittinger, K.; Volinia, S.; Caron, P.R.; Aitken, A.; [134] Kaca, W.; Kaminski, Z.J.; Kolesinska, B.; Kwinkowski, M.; Arab- Leffers, H.; Gamblin, S.J.; Smerdon, S.J.; Cantley, L.C. The struc- ski, M.; Konieczna, I. Peptides mimicking urease, methods of tural basis for 14-3-3: phosphopeptide binding specificity. Cell, manufacturing, application in diagnostic tests and the way of per- 1997, 91(7), 961-971. formance the test. Patent Applications PCT/PL2009/000106, [115] Nash, P.; Tang, X.; Orlicky, S.; Chen, Q.; Gertler, F.B.; Menden- WO/2010/071462, December 16, 2009. hall, M.D.; Sicheri, F.; Pawson, T.; Tyers, M. Multisite phosphory- [135] Kami ski, Z.J.; Paneth, P.; Rudzi ski, J. A study on the activation lation of a CDK inhibitor sets a threshold for the onset of DNA rep- of carboxylic acids by means of 2-chloro-4,6-dimethoxy-1,3,5- lication. Nature, 2001, 414(6863), 514-521. triazine and 2-chloro-4,6-diphenoxy-1,3,5-triazine. J. Org. Chem., [116] Espanel, X.; Wälchli, S.; Rückle, T.; Harrenga, A.; Huguenin- 1998, 63, 4248-4255. Reggiani, M.; van Huijsduijnen, R.H. Mapping of synergistic com- [136] Kami ski, Z.J.; Paneth, P.; O'Leary, M. Nitrogen Isotope Effects on ponents of weakly interacting protein-protein motifs using arrays of the Acylation of Aniline J. Org. Chem., 1991, 56, 5716-5719. paired peptides. J. Biol. Chem., 2003, 278(17), 15162-15167. [137] Kami ski, Z.J.; G ówka, M.L.; Olczak, A.; Martynowski, D. Ther- [117] Huang, H.; Li, L.; Wu, C.; Schibli, D.; Colwill, K.; Ma, S.; Li, C.; mal isomerization of 2- acyloxy-4,6-dimethoxy-1,3,5-triazines to 1- Roy, P.; Ho, K.; Songyang, Z.; Pawson, T.; Gao, Y.; Li, S.S.-C. acyl-3,5-dimethyl-1,3,5-triazin-2,4,6(1H,3H,5H)-triones. Crystal Defining the specificity space of the human SRC homology 2 do- structure of 1-(2,2-dimethylpropanoyloxy)3,5-dimethyl-1,3,5- main. Mol. Cell. Proteomics, 2008, 7(4), 768-784. triazin-2,4,6(1H,3H,5H)-trione. Pol. J. Chem., 1996, 70, 1316- [118] Smith, M.J.; Hardy, W.R.; Murphy, J.M.; Jones, N.; Pawson, T. 1323. Screening for PTB domain binding partners and ligand specificity [138] Kolesi ska, B.; Grabowski, S.; Konieczna, I.; Kaca, W.; Peroni, E.; using proteomederived NPXY peptide arrays. Mol. Cell Biol., Papini, A.M.; Rovero, P.; Kami ski, Z.J. In: Peptides 2006, Pro- 2006, 26(22), 8461-8474. ceedings of the 29th European Peptide Symposium, Poland, Sep- [119] Reuter, M.; Schneider-Mergener, J.; Kupper, D.; Meisel, A.; Mack- tember 3-8, 2006; Rolka, K.; Rekowski, P.; Silberring, J.; Eds., eldanz, P.; Krüger, D.H.; Schroeder, C. Regions of endonuclease Kenes International: Geneva, 2007, 616-617. EcoRII involved in DNA target recognition identified by mem- 806 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. [139] Glenska, J; Adamus-Bia ek W., Kwinkowski M., Kolesi ska B., ter pylori bound by neutralizing antibodies. Vaccine, 2010, 28, Kami ski Z., Kaca W. In: Abstracts of the VI Congress of Polish 5220-5227. Biotechnology “IV EUROBIOTECH 2011” and Central European [142] Flach, C.F.; Svensson, N.; Blomquist, M.; Ekman, A.; Raghavan, Congress of Live Science, Kraków, Poland, October 12-15, 2011; S.; Holmgren, J. A truncated form of HpaA is a promising antigen Acta Biochim. Pol., 2011; 58, Supp. 4, 67. for use in a vaccine against Helicobacter pylori. Vaccine, 2011, [140] Fujii, R.; Morihara, F.; Oku, T.; Hifumi, E.; Uda, T. Epitope map- 29(6), 1235-1241. ping and features of the epitope for monoclonal antibodies inhibit- [143] Guo, L.; Li, X.; Tang, F.; He, Y.; Xing, Y.; Deng, X.; Xi, T. Im- ing enzymatic activity of Helicobacter pylori urease. Biotechnol. munological features and the ability of inhibitory effects on enzy- Bioeng., 2004, 86(4), 434-44. matic activity of an epitope vaccine composed of cholera toxin B [141] Qiua, Y.; Wanga, Y-C.; Taoa, H-X.; Zhana, D-W.; Yuana, S-L.; subunit and B cell epitope from Helicobacter pylori urease A Wanga, P.; Wanga, L-C.; Hanb, X-P.; Li, C-S.; Li, J-K.; Liua, C-J. subunit. Appl. Microbiol. Biotechnol., 2012, 93(5), 1937-1945. Identification of B-cell epitopes in urease B subunit of Helicobac- Received: June 22, 2012 Revised: July 15, 2012 Accepted: September 03, 2012 http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Current Protein & Peptide Science Pubmed Central

Bacterial Urease and its Role in Long-Lasting Human Diseases

Current Protein & Peptide Science , Volume 13 (8) – Dec 1, 2012

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Abstract

Send Orders of Reprints at bspsaif@emirates.net.ae Current Protein and Peptide Science, 2012, 13, 789-806 789 1,* 1 1 2 Iwona Konieczna , Paulina  arnowiec , Marek Kwinkowski , Beata Kolesi ska , 2 2 1 Justyna Fr czyk , Zbigniew Kami ski and Wies aw Kaca Department of Microbiology, Institute of Biology, The Jan Kochanowski University, ul. wi tokrzyska 15, 25-406 Kielce, Poland; Technical University of Lodz, Institute of Organic Chemistry, ul. eromskiego 116, 90-924  ód , Poland. Abstract: Urease is a virulence factor found in various pathogenic bacteria. It is essential in colonization of a host organ- ism and in maintenance of bacterial cells in tissues. Due to its enzymatic activity, urease has a toxic effect on human cells. The presence of ureolytic activity is an important marker of a number of bacterial infections. Urease is also an immuno- genic protein and is recognized by antibodies present in human sera. The presence of such antibodies is connected with progress of several long-lasting diseases, like rheumatoid arthritis, atherosclerosis or urinary tract infections. In bacterial ureases, motives with a sequence and/or structure similar to human proteins may occur. This phenomenon, known as mo- lecular mimicry, leads to the appearance of autoantibodies, which take part in host molecules destruction. Detection of an- tibodies-binding motives (epitopes) in bacterial proteins is a complex process. However, organic chemistry tools, such as synthetic peptide libraries, are helpful in both, epitope mapping as well as in serologic investigations. In this review, we present a synthetic report on a molecular organization of bacterial ureases - genetic as well as structural. We characterize methods used in detecting urease and ureolytic activity, including techniques applied in disease diagnos- tic processes and in chemical synthesis of urease epitopes. The review also provides a summary of knowledge about a toxic effect of bacterial ureases on human body and about occurrence of anti-urease antibodies in long-lasting diseases. Keywords: Antibodies, long-lasting diseases, synthetic peptides, urease 1. INTRODUCTION in long-lasting diseases, like atherosclerosis or rheumatoid arthritis [6, 7]. This phenomenon will be precisely described Urease (urea amidohydrolase; EC 3.5.1.5) was the first in the next paragraphs. enzyme to be crystallized (1926). It was also the first enzy- matic protein in which the presence of nickel ions was noted 2. OCCURRENCE OF UREASE PRODUCING OR- [1]. Since then, an intensive study on urease has been con- GANISMS ducted, thanks to which a role of urease in nitrogen com- pounds circulation has been determined. It has also been Urease is produced by many different bacteria [8-17], showed that urease may be a virulence factor essential in fungi [3, 18, 19], plants [1, 3, 8, 20, 21] and even some in- various illnesses, including long-lasting diseases. vertebrates [20, 21]. Microorganisms with ureolytic proper- Urease is capable of urea hydrolysis. This compound is ties were found in soil and water as well as in human and widespread: it is found in the natural environment (water and animal bodies [8]. Ureolytic bacteria may belong to symbi- soil) and in human body, where its occurrence is connected otic natural microflora or to pathogens. In facultative anaer- with protein degradation. In humans, urea is a factor of nor- obes from intestinal microflora the level of this activity is mal functions of kidneys [2, 3]. A healthy adult excretes diverse and species characteristic [9]. about 30 g of urea per day [2]. However, it is present not Ureolytic activity is often observed in pathogenic bacte- only in urine, but also in blood serum, sweat and even in ria. Such a feature is characteristic of pathogenic Staphylo- stomach [1, 2]. Hydrolysis of urea by urease is a complex coccus strains. Over 90% of clinical methicillin resistant process. In the first step, one molecule of ammonia and one Staphylococcus aureus strains are capable of urea hydrolysis molecule of carbamate appear. In water solution, carbamate [10]. Staphylococcus leei isolated from biopsy material from spontaneously converts into the second ammonia molecule gastritis patients was also ureolytic. Uropathogenic Staphy- and carbonic acid. Next ammonia is protonated (Fig. 1). This lococcus saprophiticus is also capable of this activity [1, 11, process results in pH increase [1]. 22]. Urease is observed in Helicobacter sp., including all Urease and ammonia, generated during urea hydrolysis, Helicobacter pylori isolated from gastritis patients [1, 4, 23]. may be toxic for human tissue [4, 5] and probably have role Urease is an enzyme synthesized by pathogenic mycobacte- ria like Mycobacterium tuberculosis and Mycobacterium *Address correspondence to this author at the Department of Microbiology, bovis [12]. It was observed that anaerobic clostridia are ca- Institute of Biology, The Jan Kochanowski University, ul. wi tokrzyska pable of urea hydrolysis. About 2% of Clostridium perfring- 15, 25-406 Kielce, Poland; Tel: 48 41 349 63 05; Fax: 48 41 349 62 92; ers strains, an etiologic factor of gas gangrene, showed this E-mail: iwona.konieczna@ujk.edu.pl 1389-2037/12 $58.00+.00 © 2012 Bentham Science Publishers 790 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (1). Scheme of urea hydrolysis. feature [13]. Even some strains of Vibrio parahaemolyticus, environmental bacteria. Unique urease of Helicobacter sp. a species considered non-ureolytic, produce urease [14]. An- has a different structure. In H. pylori, urease consists of only other generally urease negative bacterial species is Es- two subunits: 26.5 kDa UreA (subunit  ) and 61.7 kDa UreB cherichia coli. Among E. coli strains, about 1% of urease- (subunit  ) coded by ureA and ureB genes [29]. A smaller positive isolates were found. This feature was connected Helicobacter sp. urease structural gene (ureA) corresponds with pathogenic O111, O157:H7, O145 and O26 enterohem- with a hypothetical fusion gene arisen from ureA and ureB orrhagic E. coli, and in O157 serogroup with sorbitol fer- typical of other bacteria, while a larger gene (ureB) is analo- menting, but non motile strains [15-17]. Proteus mirabilis is gous to ureC (Fig. 2) [30-33]. a well-known ureolytic human’s pathogen. Urease is one of Urease composed of two different polypeptides (21 kDa the major bacterial virulence factors during urinary tract in- and 65 kDa) was also identified in SL100 ureolytic coccoid fections caused by these bacteria [1, 24]. A similar phe- strain isolated from stomach biopsy material. This strain was nomenon was noted for uropathogenic Ureapasma urealyti- related to Staphylococcus cohnii and Staphylococcus xylosus, cum, Klebsiella spp., Pseudomonas spp., Corynebacterium which possess three urease subunits [34]. sp. D2, Proteus penneri, Providencia stuartii and Morgan- An active center of enzyme with two metal ions is lo- ella morganii [1, 22]. cated in the largest of structural subunits. In all ureases it is designed as UreC, except Helicobacter sp., in which case it 3. GENETIC AND STRUCTURAL ORGANIZATION is UreB [1]. Ureases are nickel-containing enzymes; how- OF BACTERIAL UREASES ever, for microaerophilic Helicobacter mustelae an iron- Urease is a nickel-containing enzyme, which requires containing urease was revealed [23]. activity of a few additional proteins for acquisition of its All bacterial ureases occur as inactive apoenzymes com- hydrolytic properties. This process involves genes coding posed of three or two types of polypeptides coded by specific structural enzyme polypeptides as well as genes coding ac- structural genes. However, additional proteins, products of cessory proteins, located in a joint cluster [1, 25]. accessory genes are required for urease activation. Those Bacterial ureases are always multimeric enzymes com- proteins (UreD, UreE, UreF, UreG and UreH) are involved posed of two or three different polypeptides [1]. In P. in transporting nickel ions into a cell and in incorporating mirabilis, three structural subunits: 11 kDa UreA (subunit  ), them into an active center of apoenzyme [35-44]. P. mirabi- 12.2 kDa UreB (subunit  ) and 61 kDa UreC (subunit  ) are lis produce active urease in presence of urea. In these bacte- found [1, 26, 27]. These polypeptides are encoded by three ria a regulatory gene ureR is present (see Fig. 2). Its product structural genes: ureA, ureB and ureC respectively [28]. is a urea inducible regulator controlling expression of re- Such organization is characteristic of most pathogenic and maining genes [1]. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 791 Fig. (2). Scheme of genetic organization of urease genes and structural composition of urease. Genetic organization of ure genes was performed basing on Microbial Genome Viever MGV 2.0 (http://mgv2.cmbi.ru.nl) for H. pylori G27, P. mirabilis HI4320, K. pneumonia 342 and Yersinia enterocolitica 8081. Under genes/polypeptides are sizes of particular genes as well as a number of amino acids of particular polypeptide were taken from NCBI database for records: CP001173 (H. pylori G27), AM942759 (P. mirabilis HI4320), NC_011283 (K. pneumonia 342) and NC_008800 (Y. enterocolitica 8081); structural genes as well as urease subunits are underlined. A highly mobile helix-turn-helix motif, located in  remaining polypeptides, similarities were smaller (Table 1.) subunit and called “flap” is essential for urease activity (see [47, 48]. Fig. 5). It may adopt two different conformations. In the In  subunit, the active center is the most conservative. “open” position, urea may enter into the active site, where Particularly stable are nickel ligands: histidines (in K. aero- hydrolyze is performed. In the “closed” position, flap covers genes His-134, His-136, His-246 and His-272), lysine (in K. the active center and blocks access to it [25]. aerogenes Lys-217) and aspartic acid (in K. aerogenes Asp- Active ureases are heterooligomeric complexes. How- 360) [1, 51]. Similarities were also observed for the flap ever, the number of particular structural subunits is always fragment from over 160 sequences of  subunit of ureases equal. In K. aerogenes urease, as well as in other tree-subunit from different microorganisms, including human pathogens. bacterial ureases, UreC/UreB/UreA molecules occur in the Many sequential identities occurred in all amino acid se- ratio 1:1:1. Likewise, for Helicobacter sp. UreB/UreA are quences. In bacteria causing human diseases, even not always in the ratio 1:1 [1]. closely related, significant conservatism was noted (Fig. 4) [52]. Urease from K. aerogenes, as well as the most of other with three active centers, one The structure of a flap region in ureases from different bacteria, is triple trimer () bacteria also possess similar conformation (Fig. 5). Bacterial in each of three  subunits. Amino- and carboxyl terminus of pathogens shows different ureolytic activity. Methods for its each subunit are free and they are able to bind additional detection, including techniques applied in disease diagnosis, compounds without disturbing the enzyme structure [1]. But are diverse. Prochrorococcus marinus sp. PCC 9511 produces urease composed according to () pattern [1, 45]. Enzymes from 5. METHODS FOR DETECTING OF UREOLYTIC Helicobacter sp. may form a more complex structure, built ACTIVITY from 12 subunits. Polypeptides  and  are linked forming trimer ( ) , where N-terminal domain of  subunit are es- Hydrolysis of urea is one of useful features in the bacte- sential in aggregation process. Then, four such trimers form ria identification. In a few infections caused by microbes, a tetrahedral complex (Fig. 3). detection of this activity is essential to disease diagnosis. Probably, such a highly complex structure of H. pylori Numerous assays are available to determine urease activ- urease enables its activity in acidic conditions, when other ity as well as to analyze kinetic behavior of urease. Most of ureases undergo nonreversible inactivation [46]. them are indirect and based on colorimetric detection of ammonia released during incubation with a buffered urea 4. CONSERVATISM OF BACTERIAL UREASES solution [53]. Ureases are considered to be conservative enzymes. One of the first methods was detecting bacterial ureolytic Among all Ure polypeptides from different bacterial species, activity based on the cultivation of microorganism on urea the highest sequence similarities were observed between containing medium (Christensen's urea medium) [54]. This is structural urease subunits from different sources. In case of the most popular qualitative method using for uropathogenes 792 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (3). Scheme of structure of bacterial ureases. Domain organization is reported for (A) H. pylori and (B) Bacillus pasteurii urease mono- mers, (C) B. pasteurii and (D) H. pylori urease trimers, and (E) H. pylori dodecamer. Table 1. The Sequence Similarity of Structural and Accessory Polypeptides of Ureases of Different Bacteria. Polypeptides Identical amino acid sequence [%] Reference Bordetella Alcaligenes K. aerogenes P. mirabilis bronchiseptica eutrophus (UreA) 84 79 73 (UreB) 63 69 67 (UreC) 70 69 68 [49] UreD 43 33 28 UreG 75 66 59 UreE 39 38 38 UreF 54 31 31 Actinobacillus Haemophilus Bacillus sp. H. pylori K. aerogenes pleuropneumoniae influenzae TB-90 (UreA) 96 67 64 57 (UreB) 86 55 53 60 (UreC) 87 62 62 66 [50] UreE 85 21 28 25 UreF 67 34 34 27 UreG 95 60 66 60 UreD 70* 24 29* 21 * - data for UreH Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 793 Fig. (4). Alignment of the amino acid sequence of urease flap fragment from pathogenic bacteria. Black - amino acid present in at least 80% of compared sequences, grey - amino acid present in at least 70% of compared sequences, white - amino acid present in less than 70% of compared sequences. Sequences from NCBI database (Accession Numbers are in figure). Alignment was performed by Clustal W 2.1 and edited with GeneDoc. Fig. (5). Conformational conservatism of bacterial ureases. All models of bacterial ureases were from ExPASy SIB Bioinformatics Resource Portal (Q7X3W5 - H. pylori; P16122 - P. hauserii; Q6GEE4 - S. aureus); overlapping was performed with RasWin Molecular Graphics Visualisation Tool (http://rasmol.org/). A - structure of H. pylori (blue) and P. hauserii (white) ureases, B - structure of H. pylori (blue) and S. aureus (white) ureases; flap region is marked by a white ellipse. like Proteus sp. In case of this bacterial species, results may most popular invasive test is a rapid urease test (RUT) that be obtained even after 4 h. A modification of Christensen requires obtaining tissue samples. However, this method is technique allows reducing assay time [54, 55]. inconvenient for patients and also incurs high costs [57, 58]. It requires biopsy specimens from defined regions of the Ureolytic activity is one of biomarkers employed to di- stomach. This material is placed on a urea-containing me- agnose H. pylori infection and to monitor bacteria eradica- dium. If bacteria are present in the specimen, the change of tion by drug treatment [56]. color resulted from alkalization of the medium is observed For diagnosis, invasive and noninvasive tests, depending [59]. A urea breath test (UBT) is commonly used among on whether endoscopy is required or not, are applied. The noninvasive tests. This method is simple, but its performance 794 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. may be slightly complicated in case of very young children urea. This property may be due to mobility of the flap region, as well as patients with certain neurological disorders [49, 50 which is different than in K. aerogenes or B. pasteurii 60]. It involves oral administration of a nontoxic isotopically ureases [46]. Due to the high activity of H. pylori urease, 14 13 labeled (C or C ) urea to a patient. Urea is hydrolyzed by local microenvironment surrounding bacterium becomes H. pylori to ammonia and isotope-containing CO . Carbon nearly neutral. Moreover, live bacterial cells adsorb on the dioxide is dissolved into blood and removed via lungs. Iso- surface enzymes released upon other H. pylori autolysis, topes are detected in exhaled air. This is a test of choice in which makes it possible for them to get to gastric mucus medical practice for detecting H. pylori infection. [61]. There layer safely [80]. Ureolytic activity is essential for surviving are also suggestions that a urea breath test may be applied for M. tuberculosis, an etiologic factor of tuberculosis, a long- diagnosing tuberculosis [62]. lasting inflammatory lung disease. Bacteria infect macro- phages. They reside in phagosome, where alkalization due to Other methods are used mainly in scientific research. ureolytic activity and subvert phagosome maturation takes Each of the methods developed for determination of ure- place. Additionally, urease activity enables bacterium to ex- olytic activity has some advantages and disadvantages (Table ist in the environment where nitrogen sources are limited to 2). urea [81]. Ureolytic activity is useful in better surviving of There are a lot of other methods based on detecting am- bacteria also in case of uropathogenes. Urease facilitates monia released by urease action, which can be determined by urinary tract infection. Infection dose for ureolytic P. mirabi- vacuum distillation, a microdiffusion, steam distillation and lis HI4320 was 1000-times lower in comparison with its electroconductivity measurement [77]. non-ureolytic mutant. Urease activity raises pH of human urine, which allows precipitation of normally soluble polyva- Fourier Transform Infrared (FTIR) spectroscopy is a lent ions to struvite and carbonate apatite. These compounds method raising a big hope for easy, quick and continuous aggregate around bacteria, forming urinary stones. Inside detection of ureolytic activity. This technique constitutes a such stones, microorganisms are protected from antibiotics radically different approach to enzymatic activity determina- and the host’s immune system [24, 83]. Urinary stones block tion. It is based on the measurement of molecular vibrations urethra or catheters leading to acute bacteriuria [24]. The energy of functional groups in organic compounds. This role of ureolytic activity in urinary stones formation was also makes FTIR spectroscopy a highly sensitive and reproduci- showed for U. urealyticum, S. saprophiticus, S. aureus and ble method. Unlike the previously discussed methods, this some Klebsiella spp., Pseudomonas spp., as well as Coryne- technique enables continuous monitoring of enzymatic reac- bacterium sp. D2, P. penneri, P. stuartii, M. morganii [1, tion by a simultaneous analysis of disappearance of substrate 22]. and the appearance of product. However, substrate as well as product must have different spectra. FTIR spectroscopy also One of the features essential in bacterial infections is enables enzyme kinetics investigation [78]. Attenuated Total persistence to the host’s cells. Schoep et al. showed that H. Reflection Fourier Transform Infrared (ATR-FTIR) spec- pylori urease have two sites (one at the N-termini of UreA troscopy was recommended to enzymatic activity analysis subunit and the other at C-termini of UreB) which were in- [79]. This technique was also applied for urease activity in- volved in persistence to endothelial cells during mouse colo- vestigations. Bands of absorbance characteristic of urea nization [29]. This observation was confirmed by investiga- (substrate) and of bicarbonate (product) could be easily tions with urease-negative H. pylori mutants incapable of monitored in time intervals (Fig. 6). [Zarnowiec et al., data colonization [1,4]. Moreover, also urease released from unpublished]. However, ATR-FTIR spectroscopy is now lyzed bacterial cells is capable of adsorption into the mucus used only in research applications, not in routine medical layer [4]. practice. Bacterial ureases affect host immune system cells. In H. pylori infection, this metalloenzyme activates monocytes and 6. UREASE AS A PATHOGENIC BACTERIA VIRU- neutrophils, which leads to secretion of inflammatory cyto- LENCE FACTOR kines and causes indirect damage to epithelial cells. Urease Bacterial ureases play a role in disease pathogenesis. is a chemotactic factor for monocytes and neutrophils. In- They are connected with urinary stones occurrence and flammatory reaction may also be initiated by adsorption of catheters blocking, pyelonephritis, ammonia encephalopathy, released enzyme into the mucus layer [4]. Induction of in- hepatic coma as well as gastritis. In many papers there are flammatory reaction was also observed for Y. enterocolitica information concerning toxic effects of bacterial ureases urease. Ability of bacterial UreB subunit to induce experi- (Table 3). mental reactive arthritis was revealed [1, 84]. The role of urease in bacterium surviving in unfavorable Urease may contribute in damaging host’s cells. Enzyme microenvironment in the host’s body is especially noticeable from H. pylori stimulates expression of inducible NO- in case of H. pylori, a causative agent of gastritis and peptic synthesizing enzyme (iNOS), which may have a cytotoxic ulceration [1, 4]. At in vitro conditions, H. pylori is sensitive effect [80]. Urease may exert a toxic effect also indirectly, to low pH. During infection, microorganisms have to pass by ammonia - the product of urea hydrolysis. During H. py- through gastric acid before reaching the protective mucus lori infection, a stimulation of an oxidative burst in neutro- layer. In these circumstances, a pathogen produces a large phils ensues and there is a release of hydrogen peroxide, amount of urease which is not observed in other bacteria which next oxidizes chlorine ions. Ammonia generated by [80]. At low pH, enzymatic activity of H. pylori urease is urease reacts with them and gives toxic monochloramine [1]. probably connected with its dodecameric structure. This en- Johnson et al. revealed, using mouse model, that ammonia zyme is also able to perform a more efficient hydrolysis of causes tissue damage also during urinary tract infections Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 795 Table 2. Characteristic of Methods for Ureolytic Activity Determination. Method Description Advantages Disadvantages Application Reference Qualitative activity is detected on the microbiological medium restricted to cultivable bacte- containing urea and phenol ria able to grow on this me- suitable for routine detection Urea-phenol facility of realization [53] red as pH indicator. Bacteria dium of activity, not recommended red-agar plate inexpensive [63] for kinetic analysis alkalize medium by hydroly- results need multiplication of sis of urea, causing change bacteria of its color. pH-dependent method. Sam- ple containing urease is electrophoresed in native allows estimation of the size equipment for electrophore- Native gel agarose or acrylamide gel. [64] of active protein sis is indispensable electrophoresis Active protein is detected [65] inexpensive time consuming after incubation of gel in solution containing urea and phenol red. Quantitative requires numerous sampling spectrophotometric method of the reaction mixture based on detection of ammo- simple [8] Phenol – sensitive to various factors very useful in full kinetic nia released during urea able to detect even a small [66] hypochlorite like temperature and time, analyses, the most frequently hydrolysis. Ammonia reacts amount of ammonia (<0.02 [67] assay pH of buffers, inhibitors used in scientific research with phenol-hypochlorite at  mol) [68] limited linearity of the cali- high pH forming indophenol bration plots needs titration with diluted HCl to determine ammonia spectrophotometric assay Nesslerization amount [8] with Nessler reagent in col- easy to perform reaction long reaction time [69] ored pH indicator solution less sensitive than phenol- hypochlorite assay spectrophotometric method GLDH has pH optimum based on coupling reaction sensitive higher than most ureases [70] Coupled of ammonia with  - alternatively, a horseradish sensitive to inhibitors [71] enzyme assay ketoglutarate in presence of peroxidase may be used for [72] difficult interpretation glutamate dehydrogenase ammonia detection expensive (GLDH) interference by potassium and other monovalent ions method of direct monitoring unaffected by inhibitors low sensitive (in ion- useful in determination of Potentiometric of ammonia ions with ion- fast in performance [73] selective electrode) the urease inhibition mecha- assays [74] selective electrodes or am- allow continuous monitoring during assay, an ionic nisms monia-selective electrode of activity strength of solution changes (there is no buffer) methods based on the urea with carbon isotope radioac- need scintillation counter useful in diagnosis of H. Isotopic [75] tive C or non-radioactive fast in performance (for C ) or mass spectrome- pylori gastric mucosa infec- methods C (there are also methods [76] 13 15 ter (for C and N ) tion based on N ). A isotope- labeled CO is detected with ureolytic P. mirabilis. In kidneys, an acute inflamma- visible [87]. Moreover, ammonia released by urease causes tion as well as necrotic cells were observed. After one week, damage to the glycosaminoglycan layer in urothelial surface, pyelonephritis was in progress. Struvite stones were noted. and disturbs its protective function [5]. After two weeks, kidneys were ulcerated and fibrosis was 796 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (6). ATR FTIR spectra of the substrate (urea), product (NaHCO ), in reaction mixture containing 0.4  g urease from Canavalia ensi- formis. Spectra of the reaction mixture were recorded at several time intervals: 0, 10, 20 and 35 min, as indicated. Table 3. Pathologic Effect of Bacterial Ureases in Human Diseases. Role of urease Bacterium species Disease Reference H. pylori gastritis, peptic ulcers [1, 4, 80] Surviving in host’s organism M. tuberculosis tuberculosis [81] E. coli hemorrhagic colitis, HUS [82] Persistence to host’s cells H. pylori gastritis, peptic ulcers [29] P. mirabilis, M. morganii, U. urealyticum Precipitation of polyvalent ions urinary tract infections [1, 22, 24, 83] and others H. pylori gastritis, peptic ulcers [4] Stimulation of inflammatory reaction Y. enterocolitica reactive arthritis [1, 84] Cytotoxic effect on host’s cells H. pylori gastritis, peptic ulcers [1, 80] Damage to glycosaminoglycan layer P. mirabilis urinary tract infections [5] Damage of tight junctions H. pylori peptic ulcers [85] Aggregation of blood platelets H. pylori gastritis, cardiovascular disease [86] HUS - hemolytic uremic syndrome Recently, a new role of H. pylori urease has been estab- lipoxygenase-mediated pathway. Such properties may have a lished. During an infection, bacteria cause increased phos- role not only in gastrointestinal, but also cardiovascular dis- phorylation of the myosin regulatory light chain. Such phos- eases [86]. phorylation regulates the function of epithelial tight junction 7. PRESENCE OF ANTI-UREASE ANTIBODIES IN complexes, which have a role in maintenance of barrier func- SERA OF PATIENTS WITH LONG-LASTING DIS- tion, cell polarity as well as intercellular adhesion. Disrup- EASES tion of tight junction is associated with a carcinogenesis process. Wroblewski et al. showed that H. pylori urease may Bacterial ureases are considered to be one of the major be connected with gastric cancer by causing damage to tight antigens in several human diseases [1, 83, 84, 88]. Hirota et junctions [85]. al. showed that this protein is immunogenic [89]. In the flap region of enzyme, the ELR motive associated with immuno- Lately, a mechanism of activating blood platelets by bac- genic antigens is present (see Fig. 4) [90]. In long-lasting terial urease has been described. Wassermann et al. showed diseases caused by ureolytic bacteria, urease may stimulate that H. pylori enzyme stimulates this process through a generation of antibodies. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 797 Infections of H. pylori are mostly chronic and, in many region of urease react with bacterial antigen. However, they cases, lifelong [91]. During a infection, an elevated level of may also recognize IKEDV motive in CCRL1 (due molecu- immunoglobulins (secretory as well as circulating) was ob- lar mimicry) and cause an inflammatory process (Fig. 7) [6]. served [88]. Different classes of antibodies were noted: in Rheumatoid arthritis (RA) is a classic long-lasting dis- the stomach - IgA and IgM, in serum - IgG and IgA. IgG ease. It is an inflammatory condition leading to joint injury. immunoglobulins remain even for a few months after bacte- During its progress, hyaline cartilages of joints as well as rium eradication [92]. Urease from this bacterium is one of bones undergo atrophy [98]. Etiology of RA is complex and, the major immunodominant antigens [93]. It is considered a despite many years of investigations, still unclear. Apart vaccine in preventing H. pylori infections. In animal model, from genetic background of RA occurrence, a role of infec- vaccination with H. pylori urease provides a significant and tious agents, like P. mirabilis, Borrelia burgdorferi, Myco- long term protection against a bacterial infection. In humans, plasma sp., M. tuberculosis, E. coli, and Porphyromonas oral administration of such a vaccine resulted in a strong gingivalis as well as some viruses was discussed [99, 100]. immune response with minimal side effects [88]. Some of them are capable of urease synthesis [12, 24]. The presence of anti-urease antibodies in H. pylori sero- Among them, the most important is P. mirabilis. Wilson et positive individuals is correlated with age and living in al. revealed a connection of bacterial urease with disease highly developed regions. Leal-Herrera, in the investigations progress. They showed a molecular mimicry between IRRET motive in P. mirabilis urease and human type XI collagen performed on a population in Mexico, revealed that the per- (LRREI sequence) present in hyaline cartilage. The observed centage of infected individuals increases with age. The pres- ence of anti-urease IgG antibodies in serum rises from less similarities concerned a sequence as well as a conformation fragments of both proteins. Simultaneously, the level of anti- than 20% in a group of individuals below 10 years old to bodies against P. mirabilis urease was significantly higher in more than 50% - in a group over 40 years old [93]. Occur- comparison to healthy individuals as well as patients with rence of anti-urease antibodies was correlated with disease ankylosing spondylitis - another autoimmune disease. Ac- severity. In patients with superficial gastritis, a low level of cording to Wilson et al. hypothesis, antibodies arising in IgG, but relatively high of IgA immunoglobulins was ob- reaction against bacterial urease function as autoantibodies served. Strong IgG reaction dominated in quiescent atrophic and recognize also human protein (collagen). This leads to gastritis individuals, whereas in patients with active atrophic primary cytotoxic damage to hyaline cartilage. In the next gastritis, reaction of IgG as well as IgA was very strong [15]. step, in an injury site the presence of cytokines, vascular Nurgalieva et al. observed the presence of IgM antibod- adhesion molecules and hydrolytic enzymes is observed. It ies, putatively recognizing a small subunit (UreA) of H. py- causes inflammation, fibrosis and destruction of joints [101]. lori urease in 94% of H. pylori-infected volunteers. The This hypothesis was confirmed in later studies. larger subunit - UreB seems to be less immunogenic. About Konieczna et al. observed for RA patient’s sera a signifi- 44% of the investigated individuals showed a positive reac- cantly higher level of antibodies recognizing synthetic pep- tion [94]. However, Burnie and Al-Dughaym showed that tide corresponding to flap epitope of P. mirabilis urease. UreB subunit of H. pylori urease has more epitopes recog- Surprisingly, they noted an elevated IgG level against pep- nized by antibodies than UreA. The level of IgG antibodies tides reflecting a sequence of flap regions from other organ- recognizing some of those epitopes was comparable with the isms (bacteria and plant). The detected antibodies also had commercial test [95]. Also in Arabski et al. study, where lower specificity. These antibodies recognized not only one levels of IgG antibodies were detected, the presence of anti- defined antigen, but also antigens with a similar sequence, bodies recognizing H. pylori UreB urease subunit was found which was probably due to instability of the immune system in almost all infected individuals. They were observed even [7]. in 70% of H. pylori negative sera. A much more interesting A role of urease in stimulation of immune response of observation was a correlation between atherosclerosis and patients with immune disease was also revealed for other the presence of anti-urease antibodies. In the investigated gram-negative bacteria. In 1993, it was showed that sera there was a significant relationship between the level of subunit of Y. enterocilitica O:3 urease is arthritogenic for antibodies bound to 8-mer synthetic peptide (which corre- rats [84]. A few years later, a high humoral response in pa- sponds to UreB minimal flap epitope of H. pylori urease) and tients with reactive arthritis triggered by Y. enterocolitica occurrence of atherosclerosis [6], an inflammatory disease O:3 was noted. IgG reacting with 19 kDa urease subunit was leading to an atheromatosus plaque in blood vessels lumen observed in over 90%, and IgA in over 50 % of investigated [96]. Earlier, Oshima et al. suggested that chronic H. pylori sera [102]. infections are connected with inflammatory processes in vessels [97]. Investigations applying synthetic peptide corre- In chronic obstructive pulmonary disease (COPD) caused sponding to 8 amino acid sequence of flap fragment of H. by nonencapsulated H. influenzae, urease is a target of hu- pylori urease revealed a similarity between this peptide and man humoral response. In almost 39% of investigated sera, a human CCRL1 (CC chemokine receptor-like 1) protein, ex- significantly higher level of antibodies reacting with bacte- pressed mainly in the heart. Based on this observation, a hy- rial urease was observed [103]. pothesis to explain a connection of H. pylori urease and athe- Anti-urease antibodies are detected even in case of rosclerosis was formulated. According to it, urease may chronic zoonosis. In patients with diagnosed brucellosis, stimulate immune system reaction during a bacterial infec- antibodies recognizing  Brucella suis urease subunit were tion. Presentation of urease fragments to Th lymphocytes detected [104]. enables synthesis of antibodies. Next, antibodies against flap 798 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (7). Molecular mimicry of flap fragment of H. pylori urease and CCRL1 and possible connection with atherosclerosis progress. In the investigations of antibodies generated as a re- sembled), apart from continuous (sequential, linear), epitopes sponse to infection, chemically defined synthetic peptides [106]. However, several strategies of epitope mapping are have a potent application. They are useful for epitope map- available. The most often used strategy is array-based oligo- ping as well as molecular mimicry studies. peptide scanning. This technique uses a library of oligo- peptide sequences from overlapping and non-overlapping 8. ORGANIC CHEMISTRY TOOLS IN IMMUNE RE- segments of a target protein and tests for their ability to bind SPONSE INVESTIGATIONS the antibody of interest. This method is fast and relatively inexpensive, and specifically suited to profile epitopes for a Organic chemistry enables constructing several new tools large number of candidate antibodies against defined targets for investigations of response of the immune system against [108]. infectious agents. One of these tools is a library of synthetic peptides with a chemically defined sequence. Such libraries So far the most general approach for epitope mapping has are used for detecting antibodies as well as for estimating been developed by Ronald Frank [109]. The applied meth- their variety and specificity [105]. Synthetic peptide libraries odology, known as SPOT synthesis, is a special type of solid also provide epitope mapping of protein antigens, the process phase peptide synthesis proceeding directly on the membrane of locating the epitope on the protein surface or in the protein support, inside relatively small, separated spots regarded as sequence [106]. There are many peptide synthesis methods: separate reaction vessels. The method was initiated as an biological (peptide is expressed on the surface of bacterium uncomplicated manual technique for parallel chemical syn- or phage) or fully synthetic (peptide is synthesized on the thesis of peptide arrays followed by an assay with appropri- abiotic surface like cellulose or polypropylene) [107]. ate interacting molecules performed directly on the mem- brane or in solution (Fig. 8) [105]. Determination of antigenic determinants of protein may be intricate considering the existence of discontinuous (as- Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 799 Fig. (8). Scheme of SPOT synthesis. SPOT method is useful for mapping not only linear, but Synthesis of peptide on the membrane surface allows also discontinuous epitopes as well as for characterizing an- application of simple techniques, like dot-blot, for further tibodies [110, 111, 112]. It was even applied for identifica- investigations of e.g. bounded antibodies. However, the re- tion of peptide mimicking the structure of an epitope (mimo- sults obtained strongly depend on the structure of the linker tope) [110]. A broad variety of other biomolecular binding fragment. The most classical linkers are prepared using 1-3 events or enzymatic modifications can be investigated by residues of  -alanine or glycine to separate peptide assem- using peptide arrays (prepared by the SPOT technique) such bling with biomolecules used in assay out of the membrane as protein-protein interactions [113-118] protein-DNA inter- [109, 128]. In SPOT technology, different linkers are also actions [119] peptide-cell interactions [120-122] or enzyme- applied (like Carboxy-Frank-Linker, p-hydroxy- substrate interactions [123-127]. methylbenzoic acid (HMB) linker, the Rink-amide linker or 800 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. Fig. (9). Synthesis of peptides with free N-termini anchored by isocyanuric linker. 4-hydroxymethyl-phenoxy acetic acid (HMPA) and 4-(4- A different type of anchoring the peptide chain to cellu- hydroxymethyl-3-methoxyphenoxy)-butyric acid (HMPB) lose matrix was proposed by Kaminski and co-workers [6, linkers) enabling cleavage of peptides from the support [129- 134-136]. 1-Acyl-3,5-dimethyl-1,3, 5-triazin-2,4,6 (1H,3H, 133]. 5H)-trion derivatives serve both as a spacer and a linker. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 801 This isocyanuric linker has been introduced by thermal iso- on the synthesis of the target amino acid sequence as a series merization of 2-acyloxy-4,6-dimethoxy-1,3,5-triazines al- of overlapping peptides on polypropylene pins [30]. ready immobilized on the cellulose support [137]. A syn- Studies on urease epitopes were also performed using thetic procedure leading to peptides anchored to cellulose by other techniques. One of the most interesting new develop- 1-acyl-3,5-dimethyl-1,3,5-triazin-2,4,6(1H,3H,5H)-trion ments in the search for novel antigens applied a computa- (iso-MT) is shown in (Fig. 9). In the first step, chloro- tional method to predict T-cell epitopes using the whole ge- triazine immobilized on cellulose was treated with N- nome sequence information [128]. methylmorpholine yielding N-triazinylammonium chloride, In a more classical approach, antibody binding epitopes which is activating the carboxylic function of Fmoc- in H. pylori urease were determined with monoclonal anti- protected amino acid yielding superactive triazine type ester [135, 136], which, in refluxing toluene, rearranges to a stable bodies produced in Balb/c mice. After digestion of bacterial isocyanuric derivative. urease with trypsin followed by separation of peptides with affinity chromatography, monoclonal antibodies were ap- During peptide synthesis, anchoring method is essential plied to identify epitopes. Next, amino acid sequences of for further reaction of peptide and antibody (Table 4). isolated peptides were determined by mass spectroscopic For isocyanuric linker, interactions with antibodies were analysis. Fujii et al. showed that two such peptides, with the found more selective [138]. This linker was applied during sequence SVELIDIGGNRRIFGFNALVDR and IFGFNAL synthesis of peptides corresponding to a flap region of dif- VDR, were recognized by two monoclonal antibodies ferent ureases (bacterial and plant) based on the flap region (MAb): HpU-2 and HpU-18 respectively. Based on the data from H. pylori urease. Those peptides were useful in investi- of competitive binding determined by using surface plasmon gating human sera. It was possible to differentiate sera of resonance and analysis of the epitope for HpU-2 and HpU- patients with autoimmune diseases like rheumatoid arthritis 18, it has been found that both MAb recognize almost the or atherosclerosis from sera of healthy donors [6, 7]. On the same position on the UreA subunit of H. pylori urease. An other hand, a library of 361 peptides, where each peptide had unanticipated result was suppressing of urease activity via an a sequence with one substituted amino acids in a defined allosteric effect, which might cause a distortion of the con- position, was applied to determine amino acids which are formation of the enzyme. On the other hand, the second essential for antibody binding [139]. mechanism observed in the case of several MAb studied For epitope mapping of UreA and UreB subunits of H. (which in fact possessed a weaker inhibitory effect, such as pylori urease, Geysen’s method was adopted, which is based HpU-17 and -20) is assumed that MAb B binds to the vicin- Table 4. Interaction of Antibodies with H. pylori Urease Epitopes with Free N-Termini Anchored on Cellulose. Epitope Peptide sequence Reaction Specificity cellulose H C N (Aaa) -NH n 2 HN-CHHLDKSIKEDVQFADSRI-COO-cellulose - UB-33 H N-CHHLDKSIKEDVQFADSRI--Ala-COO-cellulose - 0% H N-CHHLDKSIKEDVQFADSRI- -Ala- -Ala--Ala-COO-cellulose - cellulose N N Me (Aaa) -NH n 2 H N-CHHLDKSIKEDVQFADSRI--Ala-iso-MT-cellulose +, m UB-33 H2N-CHHLDKSIKEDVQFADSRI-iso-MT-cellulose +, s 100% H N-SIKEDVQF--Ala-iso-MT-cellulose +, s F-8 HN-SIKEDVQF-iso-MT-cellulose +, m a) - (no reaction); + (reaction); s (strong); m (medium). 802 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. ity of the active site, resulting in the reduction of the urease Urease, although investigated for a long time, still seems to be an unexplored enzyme. activity [140]. Molecular biology methods are also used in H. pylori CONFLICT OF INTEREST urease epitope determination. Nineteen truncated fragments of gene coding UreB subunit were amplified and cloned into The author(s) confirm that this article content has no con- the prokaryotic expression vector pET-28a (+) or pGEX-4T- flicts of interest. 2. After verification, the constructs obtained were trans- ACKNOWLEDGEMENTS formed into Escherichia coli which expressed recombinant proteins. Using three MAbs against UreB of H. pylori This work was supported by grant N N304 044639 from (A1H10, A3C10, and B3D9) three linear B-cell epitopes, Ministry of Sciences and High Education (Poland). probably useful as the targets for development of epitope- based vaccines against H. pylori, were identified. These epi- REFERENCES topes were localized in the aa regions: 158-172 [1] Mobley, H.L.T.; Island, M.D.; Hausinger, R.P. Molecular biology (GGGTGPADGTNATTI), 181-195 (WMLRAAEEYS of microbial ureases. Microbiol. Rev., 1995, 59(3), 451-480. MNLGF), and 349-363 (TLHDMGIFSITSSDS) of UreB [2] Newsholme, E.; Leech, A. Functional Biochemistry in Health and nd Disease, 2 ed.; John Wiley & Sons, Chichester, 2011. [141]. [3] Sirko, A.; Brodzik, R. Plant ureases: Roles and regulation. Acta Urease is highly expressed by all strains of H. pylori and is Biochim. Pol., 2000, 47(4), 1189-1195. immunogenic. Additionally this enzyme could stimulate [4] Dunn, B.E.; Phadnis S.H. Structure, Function and Localization of Helicobacter pylori Urease. Yale J. Biol. Med., 1998, 71(2), 63-73. generation of antibodies able to inhibit its activity. For this [5] Follmer, C. Ureases as a target for the treatment of gastric and reason, it seems to be a promising vaccine target. However, urinary infections. J. Clin. Pathol., 2010, 63(5), 424-430. vaccination of urease may not give a sufficient protective [6] Arabski, M.; Konieczna, I.; So owiej, D.; Rogo , A.; Kolesi ska, effect. A combination of urease with other antigens may B.; Kami ski, Z.; Kaca, W. Are anti-H. pylori urease antibodies in- yield better results. It seems that a fusion of UreB urease volved in atherosclerotic disease? Clin. Biochem., 2010, 43(1-2), subunit with truncated HpaA surface protein may give a bet- 115-123. [7] Konieczna, I.; Kwinkowski, M.; Kolesi ska, B.; Kami ski, Z.; ter protection than either protein alone [142]. Fr czyk, J.; arnowiec, P.; Kaca, W. Detection of Antibodies There were also attempts to design a vaccine using mu- against Synthetic Peptides Mimicking Ureases Fragments in Sera cosal adjuvant cholera toxin B subunit (CTB) and an epitope of Rheumatoid Arthritis Patients. Prot. Pept. Lett., 2012, 19(11), 1149-1154. (UreA 183-203) of H. pylori urease. Both peptides were [8] Mobley, H.L.T.; Hausinger, R.P. Microbial ureases: significance, bound with the linker (DPRVPSS) to avoid the formation of regulation, and molecular characterization. Microbiol. Rev., 1989, new epitopes. The CTB-UreA epitope vaccine had good im- 53(1), 85-108. munogenicity and immunoreactivity and induced specific [9] Suzuki, K.; Benno, Y.; Mitsuoka, T.; Takebe, S.; Kobashi, K.; neutralizing antibodies which showed an effectively inhibi- Hase, J. Urease-Producing Species of Intestinal Anaerobes and tory effect on H. pylori urease enzymatic activity [143]. Their Activities. Appl. Environ. Microbiol., 1979, 37(3), 379-382. [10] Murchan, S.; Aucken, H.M.; O’Neill, G.L.; Ganner, M.; Cookson, Today, experiments to identify and characterize linear B.D. Emergence, Spread, and Characterization of Phage Variants antibody epitopes using peptide scans, amino acids scans, of Epidemic Methicillin-Resistant Staphylococcus aureus 16 in substitutional analyses, truncation libraries, deletion librar- England and Wales. J. Clin. Microbiol., 2004 42(11), 5154-5160. [11] Jin, M.; Rosario, W.; Watler, E.; Calhoun, D.H. Development of a ies, cyclization scans, all types of combinatorial libraries and large-scale HPLC-based purification for the urease from Staphylo- randomly generated libraries of single peptides are standard coccus leei and determination of subunit structure. Protein Expr. techniques widely applied even in non-specialized laborato- Purif. 2004, 34(1), 111-117. ries [135]. [12] Clemens, D.L.; Lee, B-Y.; Horwitz, M.A. Purification, Characteri- zation, and Genetic Analysis of Mycobacterium tuberculosis CONCLUSIONS Urease, a Potentially Critical Determinant of Host-Pathogen Inter- action. J. Bacteriol., 1995, 177(19), 5644-5652. Urease is an enzyme studied for a long time. Its structure, [13] Dupuy, B.; Daube, G.; Popoff, M.R.; Cole, S.T. Clostridium per- fringens Urease Genes Are Plasmid Borne. Infect. Immun., 1997, synthesis and biochemical activity are known. There are also 65(6), 2313-2320. many studies concerning urease toxic effect on human tis- [14] Lam, S.; Yeo, M. Urease-Positive Vibrio parahaemolyticus Strain. sues. However, its role in long-lasting autoimmune diseases J. Clinical Microbiol., 1980, 12(1), 57-59. is still controversial. Nevertheless, the presence of molecular [15] Futagami, S.; Takahashi, H.; Norose, Y.; Kobayashi, M. Systemic mimicry between bacterial ureases and human proteins has and local immune responses against Helicobacter pylori urease in patients with chronic gastritis: distinct IgA and IgG productive been suggested [7, 101]. Proteins containing motives, similar sites. Gut, 1998, 43(2), 168-175. to infectious agents, may function as autoantigens. In de- [16] Nakano, M.; Iida, T.; Ohnishi, M.; Kurokawa, K.; Takahashi, A.; scribed autoantigenes, some similarities to ureases may be Tsukamoto, T.; Yasunaga, T.; Hayashi, T.; Honda, T. Association found [90, 101]. It was proved that this enzyme stimulates of the Urease Gene with Enterohemorrhagic Escherichia coli antibodies synthesis [89], but determination of epitopes in Strains Irrespective of Their Serogroups. J. Clin. Microbiol., 2001, urease protein may be difficult and non-conclusive. There- 39(12), 4541-4543. [17] Orth, D.; Grif, K.; Dierich, M.P.; Würzner, R. Prevalence, structure fore investigations applying synthetic peptides could be very and expression of urease genes in Shiga toxin-producing Es- helpful in mapping epitopes both in infectious agents pro- cherichia coli from humans and the environment. Int. J. Hyg. Envi- teins as well as in determining amino acids located in epi- ron.-Health, 2006, 209(6), 513-520. topes which are essential for human humoral response [139]. [18] Tange, Y.; Niwa, O. Identification of the ure1+ gene encoding urease in fission yeast. Curr. Genet., 1997, 32(3), 244-246. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 803 [19] Yu, J.J.; Smithson, S.L.; Thomas, P.W.; Kirkland, T.N.; Cole, G.T. that are competent for in vivo enzyme activation. J. Bacteriol., Isolation and characterization of the urease gene (URE) from the 1995, 177(8), 1947-1951. pathogenic fungus Coccidioides immitis. Gene, 1997, 198(1-2), [40] Sriwanthana, B.; Island, M.D.; Maneval, D.; Mobley, H.L.T. Single 387-391. step purification of Proteus mirabilis urease accessory protein [20] Zonia, L.E.; Stebbins, N.E.; Polacco, J.C. Essential role of urease UreE, a protein with a naturally occurring histidine tail, by nickel in germination of nitrogen-limited Arabidopsis thaliana seeds. chelate affinity chromatography. J. Bacteriol., 1994, 176(22), Plant Physiol., 1995, 107(4), 1097-1103. 6836-6841. [21] Pedrozo, H.A.; Schwartz, Z.; Luther, M.; Dean, D.D.; Boyan, B.D.; [41] Sriwanthana, B.; Island, M.D.; Mobley, H.L.T. Sequence of the Wiederhold, M.L. A mechanism of adaptation to hypergravity in Proteus mirabilis urease accessory gene ureG. Gene, 1993, 129(1), the statocyst of Aplysia californica. Hear Res., 1996, 102(1-2), 51- 103-106. 62. [42] Voland, P.; Weeks, D.L.; Marcus, E.A.; Prinz, C.; Sachs, G.; Scott, [22] Hedelin, H. Uropathogens and urinary tract concretion formation D. Interactions among the seven Helicobacter pylori proteins en- and catheter encrustations. Int. J. Antimicrob. Agents, 2002, 19(6), coded by the urease gene cluster. Am. J. Physiol. Gastrointest. 484-487. Liver Physiol., 2003, 284(1), 96-106. [23] Carter, E.L.; Proshlyakov, D.A.; Hausinger, R.P. Apoprotein isola- [43] Park, I.-S.; Hausinger, R.P. Requirement of CO2 for in vitro assem- tion and activation, and vibrational structure of the Helicobacter bly of the urease nickel metallocenter. Science, 1995, 267(5201), mustelae iron urease. J. Inorg. Biochem., 2011, 111, 195-202. 1156-1158. [24] Rózalski, A.; Kwil, I.; Torzewska, A.; Baranowska, M.; Staczek, P. [44] Musiani, F.; Zambelli, B.; Stola, M.; Ciurli, S. Nickel trafficking: Proteus bacilli: features and virulence factors. Postepy Hig. Med. insights into the fold and function of UreE, a urease metallochaper- Dosw., 2007, 61, 204-219. one. J. Inorg. Biochem., 2004, 98(5), 803-813. [25] Lv, J.; Jiang, Y.; Yu, Q.; Lu, S. Structural and functional role of [45] Palinska, K.A.; Jahns, T.; Rippka, R.; Tandeau de Marsac, N. Pro- nickel ions in urease by molecular dynamics simulation. J Biol chlorococcus marinus strain PCC 9511, a picoplanktonic cyano- Inorg Chem., 2011, 16(1), 125-135. bacterium, synthesizes the smallest urease. Microbiology, 2000, [26] Mörsdorf, G.; Kaltwasser, H. Cloning of the genes encoding urease 146, 3099-3107. from Proteus vulgaris and sequencing of the structural genes. [46] Ha, N.-C.; Oh, S-T.; Sung, J.Y.; Cha, K.A.; Lee, M.H.; Oh, B.-H. FEMS Microbiol. Lett., 1990, 54(1-3), 67-73. Supramolecular assembly and acid resistance of Helicobacter py- [27] Walz, S.E.; Wray, S.K.; Hull, S.I.; Hull, R.E. Multiple proteins lori urease. Nat. Struct. Biol., 2001, 8, 505-509. encoded within the urease gene complex of Proteus mirabilis. J. [47] McMillan, D.J.; Mau, M.; Walker, M.J. Characterisation of the Bacteriol., 1988, 170(3), 1027-1033. urease gene cluster in Bordetella bronchiseptica. Gene, 1998, [28] Jones, B.D.; Mobley, H.L.T. Proteus mirabilis urease: nucleotide 208(2), 243-251. sequence determination and comparison with jack bean urease. J. [48] Bosse, J.T.; MacInnes, J.I. Genetic and Biochemical Analyses of Bacteriol., 1989, 171(12), 6414-6422. Actinobacillus pleuropneumoniae Urease. Infect. Immun., 1997, [29] Schoep, T.D.; Fulurija, A.; Good, F.; Lu, W.; Himbeck, RP, 65(11), 4389-4394. Schwan C.; Choi S. S.; Berg D.E.; Mittl P.R.E.; Benghezal M.; [49] Monteiro, L.; de Mascarel, A.; Sarrasqueta, A.M.; Bergey, B.; Marshall B.J. Surface Properties of Helicobacter pylori Urease Barberis, C.; Talby, P.; Roux, D.; Shouler, L.; Goldfain, D.; Complex Are Essential for Persistence. PLoS ONE, 2010, 5(11), Lamouliatte, H.; Megraud, F. Diagnosis of Helicobacter pylori in- e15042. doi:10.1371/journal.pone.0015042 fection: noninvasive methods compared to invasive methods and [30] Clayton, C.L.; Pallen, M.J.; Kleanthous, H.; Wren, B.W.; Ta- evaluation of two new tests. Am. J. Gastroenterol., 2001, 96(2), baqchali, S. Nucleotide sequence of two genes from Helicobacter 353-358. pylori encoding for urease subunits. Nucleic Acids Res., 1990, [50] Falsafi, T.; Favaedi, R.; Mahjoub, F.; Najafi M. Application of 18(2), 362. Stool-PCR test for diagnosis of Helicobacter pylori infection in [31] Ferrero, R.L.; Labigne, A. Cloning, expression and sequencing of children World J. Gastroenterol., 2009, 15(4), 484-488. Helicobacter felis urease genes. Mol. Microbiol., 1993, 9(2), 323- [51] Ragsdale, S.W. Nickel-based Enzyme Systems. J. Biol. Chem., 333. 2009, 284(28), 18571-18575. [32] Labigne, A.; Cussac, V.; Courcoux, P. Shuttle cloning and nucleo- [52] Konieczna, I. Characteristic of the bacterial ureases molecular tide sequences of Helicobacter pylori genes responsible for urease variety and estimation of reactivity of the human anti-urease anti- activity. J. Bacteriol., 1991, 173(6), 1920-1931. bodies. PhD Thesis, The Jan Kochanowski University of Humani- [33] Solnick, J.V.; O’Rourke, J.; Lee, A.; Tompkins, L.S. Molecular ties and Sciences: Kielce, April, 2010. analysis of urease genes from a newly identified uncultured species [53] Hamilton-Miller, J.M.T.; Gargan, R.A. Rapid screening for urease of Helicobacter. Infect. Immun., 1994, 62(5), 1631-1638. inhibitors. Invest. Urol., 1979, 16(5), 327-328. [34] Lee, S.G.; Calhoun, D.H. Urease from a Potentially Pathogenic [54] Christensen, W.B. Urea decomposition as a means of differentiat- Coccoid Isolate: Purification, Characterization, and Comparison to ing Proteus and paracolon cultures from each other and from Sal- Other Microbial Ureases. Infect. Immun., 1997, 65(10), 3991-3996. monella and Shigella types. J. Bacteriol., 1946, 52(4), 461-466. [35] Lee, M.H.; Mulrooney, S.B.; Renner, M.J.; Markowicz, Y.; Haus- [55] Hussain Qadri, S.M.; Zubairi, S.; Hawley, H.P.; Mazlaghani, H.H.; inger, R.P. Klebsiella aerogenes urease gene cluster: sequence of Ramirez, E.G. Rapid test for determination of urea hydrolysis. An- ureD and demonstration that four accessory genes (ureD, ureE, tonie van Leeuwenhoek, 1984, 50(4), 417-423. ureF, ureG) are involved in nickel metallocenter biosynthesis. J. [56] Bell, G.D.; Weil, J.; Harrison, G.; Morden, A.; Jones, P.H.; Gant, Bacteriol., 1992, 174(13), 4324-4330. P.W.; Trowell, J.E.; Yoong, A.K.; Daneshmend, T.K.; Logan, [36] Lee, M.H.; Pankratz, H.S.; Wang, S.; Scott, R.A.; Finnegan, M.G.; R.F.A. 14C-urea breath analysis, a non-invasive test for Campy- Johnson, M.K.; Ippolito, J.A.; Christianson, D.W.; Hausinger, R.P. lobacter pylori in the stomach. Lancet, 1987, 329(8546), 1367- Purification and characterization of Klebsiella aerogenes UreE pro- 1368. tein: a nickel-binding protein that functions in urease metallocenter [57] Vakil, N.; Rhew, D.; Soll, A.; Ofman, J.J. The costeffectiveness of assembly. Protein Sci., 1993, 2(6), 1042-1052. diagnostic testing strategies for Helicobacter pylori. Am. J. Gastro- [37] Mulrooney, S.B.; Hausinger, R.P. Sequence of the Klebsiella aero- enterol., 2000, 95(7), 1691-1698. genes urease genes and evidence for accessory proteins facilitating [58] Mansour-Ghanaei, F.; Sanaei, O.; Joukar, F. Clinical Validation of nickel incorporation. J. Bacteriol., 1990, 172(10), 5837-5843. an Office-Based 14C-UBT (Heliprobe) for H. pylori Diagnosis in [38] Park, I.-S.; Carr, M.B.; Hausinger, R.P. In vitro activation of urease Iranian Dyspeptic Patients. Gastroenterol. Res. Pract., 2011, 2011, apoprotein and role of UreD as a chaperone required for nickel 930941. metallocenter assembly. Proc. Natl. Acad. Sci. USA, 1994, 91(8), [59] Foroutan, M.; Loloei, B.; Irvani, S.; Azargashb, E. Accuracy of 3233-3237. rapid urease test in diagnosing Helicobacter pylori infection in pa- [39] Park, I.-S.; Hausinger, R.P. Evidence for the presence of urease tients using NSAIDs. Saudi. J. Gastroenterol., 2010, 16(2), 110- apoprotein complexes containing UreD, UreF, and UreG in cells 112. 804 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. [60] Krogfelt, K.A.; Lehours, P.; Megraud, F. Diagnosis of Helicobac- [83] Coker, C.; Poore, C.A.; Li, X.; Mobley, H.L.T. Pathogenesis of ter pylori Infection. Helicobacter, 2005, 10(Suppl 1), 5-13. Proteus mirabilis urinary tract infection. Microbes Infect., 2000, [61] Artiko, V.M.; Obradovi , V.B.; Petrovi , N.S.; Davidovi , B.M.; 2(12), 1497-1505. Gruji -Adanja, G.S.; Nasti -Miri , D.R.; Milosavljevi . T.N. 14C- [84] Probst, P.; Hermann, E.; Meyer Zum Buschenfelde, K.-H.; urea breath test in the detection of Helicobacter pylori infection. Fleischer, B. Identification of the Yersinia enterocolitica Urease Nuc. Med. Rev., 2001, 4(2), 101-103. Subunit as a Target Antigen for Human Synovial T Lymphocytes [62] Jassal, M.S.; Nedeltchev, G.G.; Lee, J-H.; Choi, S.W.; Atudorei, in Reactive Arthritis. Infect. Immun., 1993, 61(10), 4507-4509. V.; Sharp, Z.D.; Deretic, V.; Timmins, G.S.; Bishai, WR. [C]- [85] Wroblewski, L.E.; Shen, L.; Ogden, S.; Romero-Gallo, J.; Lapierre, Urea Breath Test as a Novel Point-of-Care Biomarker for Tubercu- L.A.; Israel, D.A.; Turner, J.R.; Peek, R.M.Jr. Helicobacter pylori losis Treatment and Diagnosis. PLoS ONE, 2010, 5(8), e12451. dysregulation of gastric epithelial tight junctions by urease- [63] Ruiz-Herrera, J.; Gonzalez, J. A continuous method for the meas- mediated myosin II activation. Gastroenterology, 2009, 136(1), urement of urease activity. Anal. Biochem., 1969, 31, 366-374. 236-246. [64] Blattler, D.P.; Contaxis, C.C.; Reithel, F.J. Dissociation of urease [86] Wassermann, G.E.; Olivera-Severo, D.; Uberti, A.F.; Carlini, C.R. by glycol and glycerol. Nature, 1967, 216, 274-275. Helicobacter pylori urease activates blood platelets through a [65] Shaik-M, M.B.; Guy, A.L.; Pancholy, S.K. An improved method lipoxygenase-mediated pathway. J. Cell. Mol. Med., 2010, 14(7), for the detection and preservation of urease activity in polyacryla- 2025-2034. mide gel. Anal. Biochem., 1980, 103, 140-143. [87] Johnson, D.E.; Russell, R.G.; Lockatell, C.V.; Zulty, J.C.; Warren, [66] Weatherburn, M.W. Phenol-hypochlorite reaction for determination J.W.; Mobley, H.L.T. Contribution of Proteus mirabilis Urease to of ammonia. Anal. Chem., 1967, 39, 971-974. Persistence, Urolithiasis, and Acute Pyelonephritis in a Mouse [67] Krajewska, B.; Chudy, M.; Drozdek, M.; Brzozka, Z. Potentiomet- Model of Ascending Urinary Tract Infection Infect. Immun., 1993, ric Study of Urease Kinetics over pH 5.36 -8.21. Electroanalysis, 61(7), 2748-2754. 2003, 15(5-6), 460-466. [88] Burne R.A.; Chen, Yi-Y.M. Bacterial ureases in infectious dis- [68] Juszkiewicz, A.; Kot, M.; Zaborska, W. Calorimetric study of eases. Microbes Infect., 2000, 2(5), 533-542. inhibition of urease by 2-mercaptoethanol. Procedures based upon [89] Hirota, K.; Nagata, K.; Norose, Y.; Futagami, S.; Nakagawa, Y.; integrated rate equations. Thermochim. Acta, 1998, 320(1-2), 45- Senpuku, H.; Kobayashi, M.; Takahashi, H. Identification of an an- 52. tigenic epitope in Helicobacter pylori urease that induces neutraliz- [69] Lin, Y-L.; Chen, Ch-T.; Lin, S-C.; Lee, C.; Kuo, H-S.; Shih, C-M.; ing antibody production. Infect. Immun., 2001, 69(11), 6597-6603. Hsu, Y-H.; Chin, Y-P.; Chan, E-C. A simple method to determine [90] Backes, C.; Ludwig, N.; Leidinger, P.; Harz, C.; Hoffmann, J.; urea concentration using intact Helicobacter pylori and Bromo Keller, A.; Meese, E.; Lenhof, H-P. Immunogenicity of autoanti- Cresol Purple as a pH indicator. Biotechnol. Lett., 2000, 22(13), gens. BMC Genom., 2011, 12, 340. 1077-1079. [91] Thomas, J.E.; Whatmore, A.M.; Barer, M.R.; Eastham, E.J.; Ke- [70] Kaltwasser, H.; Schlegel, H.G.. NADH-dependent coupled assay hoe, M.A. Serodiagnosis of Helicobacter pylori Infection in Child- for urease and other ammonia-producing systems. Anal. Biochem., hood. J. Clin. Microbiol., 1990, 28(12), 2641-2646. 1966, 16(1), 132-138. [92] Dunn, B.E.; Cohen, H.; Blaser, M.J. Helicobacter pylori. Clin. [71] Gambhir, A.; Gerard, M.; Mulchandani, A.K.; Malhotra, B.D. Microbiol. Rev., 1997, 10(4), 720-741. Coimmobilization of Urease and Glutamate Dehydrogenase in [93] Leal-Herrera, Y.; Torres, J.; Perez-Perez, G.; Gomez, A.; Monath, Electrochemically Prepared Polypyrrole-Polyvinyl Sulfonate Films. T.; Tapia-Conyer, R.; Muñoz, O. Serologic IgG Response To Appl. Biochem. Biotechnol., 2001, 96(1-3), 249-257. Urease In Helicobacter pylori-Infected Persons From Mexico Am. [72] Stutts, P.; Fridovich, I. A continual spectrophotometric determina- J. Trop. Med. Hyg., 1999, 60(4), 587-592. tion of ammonia-producing systems. Anal. Biochem., 1964. 8, 70- [94] Nurgalieva, Z.Z.; Conner, M.E.; Opekun, A.R.; Zheng, C.Q.; 74. Elliott, S.N.; Ernst, P.B.; Osato, M.; Estes, M.K.; Graham, D.Y. B- [73] Katz, S.A. Direct potentiometric determination of urease activity. Cell and T-Cell Immune Responses to Experimental Helicobacter Anal. Chem., 1964, 36(13), 2500-2501. pylori Infection in Humans. Infect. Immun., 2005, 73(5), 2999- [74] Montalvo, J.C., Jr. An improved urease electrode. Anal. Biochem., 3006. 1970, 36(2), 357-363. [95] Burnie, J.P.; Al-Dughaym, A. The application of epitope mapping [75] McDonald, J.A.; Speeg, K.V.; Campbell, J.W. Urease: a sensitive in the development of a new serological test for Helicobacter pylori and specific radiometric assay. Enzymologia, 1972, 42, 1-9. infection. J. Immunol. Methods, 1996, 194(1), 85-94. [76] Wrong, O.M.; Vince, A.J.; Waterlow, J.C. The contribution of [96] Milioti, N.; Bermudez-Fajardo, A.; Penichet, M.L.; Ernesto, O.-O. endogenous urea to face alammonia in man, determined by 15N- Antigen-Induced Immunomodulation in the Pathogenesis of Athe- labeling of plasma urea. Clin. Sci., 1985, 68(2), 193-199. rosclerosis. Clin. Dev. Immunol., 2008, 2008, 723539. doi: [77] Solomon, C.M.; Alexander, J.A.; Glibert P.M. Measuring urease 10.1155/2008/723539. activity in aquatic environmental samples. Limnol. Oceanogr. [97] Oshima, T.; Ozono, R.; Yano, Y.; Oishi, Y.; Teragawa, H.; Higa- Methods, 2007, 5, 280-288. shi, Y.; Yoshizumi, M.; Kambe, M. Association of Helicobacter [78] Karmali, K.; Karmali, A.; Teixeira, A.; Curto, M.J. The use of pylori Infection With Systemic Inflammation and Endothelial Dys- Fourier transform infrared spectroscopy to assay for urease from function in Healthy Male Subjects. J. Am. Coll. Cardiol., 2005, Pseudomonas aeruginosa and Canavalia ensiformis. Anal. Bio- 45(8), 1219-1222. chem., 2004, 331(1), 115-121. [98] Aletaha, D.; Neogi, T.; Silman, A.J.; Funovits, J.; Felson, D.T.; [79] Kumar, S.; Barth, A. Following Enzyme Activity with Infrared Bingham, C.O.; Birnbaum, N.S.; Burmester, G.R.; Bykerk, V.P.; Spectroscopy. Sensors, 2010, 10, 2626-2637. Cohen, M.D.; Combe, B.; Costenbader, K.H.; Dougados, M.; Em- [80] Olivera-Severo, D.; Wassermann1, G.E.; Carlini, C.R. Ureases ery, P.; Ferraccioli, G.; Hazes. J.M.W.; Hobbs, K.; Huizinga, display biological effects independent of enzymatic activity. Is T.W.J.; Kavanaugh, A.; Kay, J.; Kvien, T.K.; Laing, T.; Mease, P.; there a connection to diseases caused by urease-producing bacteria? Menard, H.A.; Moreland, L.W.; Naden, R.L.; Pincus, T.; Smolen, Braz. J. Med. Biol. Res., 2006, 39(7), 851-861. J.S.; Stanislawska-Biernat, E.; Symmons, D.; Tak, P.P.; Upchurch, [81] Lin, W.; Mathys, V.; Ang, E.L.; Koh, V.H.; Martínez Gómez, J.M.; K.S.; Vencovsk , J.; Wolfe, F.; Hawker, G. 2010 Rheumatoid Ar- Ang, M.L.; Zainul Rahim, S.Z.; Tan, M.P.; Pethe, K.; Alonso, S. thritis Classification Criteria. Arthritis & Rheumatism, 2010, 62(9), Urease activity represents an alternative pathway for Mycobacte- 2569-2581. rium tuberculosis nitrogen metabolism. Infect. Immun., 2012, [99] Rashid, T.; Ebringer, A. Rheumatoid arthritis is linked to Pro- 80(8), 2771-2779. teus—the evidence. Clin. Rheumatol., 2007, 26(7), 1036-1043. [82] Steyert, S.R.; Kaper, J.B. Contribution of Urease to Colonization [100] Tobón, G.J.; Youinou, P.; Saraux, A. The environment, geo- by Shiga Toxin-Producing Escherichia coli. Infect. Immun., 2012, epidemiology, and autoimmune disease: Rheumatoid arthritis. J. 80(8):2589-2600. Autoimmun. 2010, 35(1), 10-14. Bacterial Urease and its Role in Long-Lasting Human Diseases Current Protein and Peptide Science, 2012, Vol. 13, No. 8 805 [101] Wilson, C.; Tiwana, H.; Ebringer, A. Molecular mimicry between brane-bound peptide repertoires. J. Biol. Chem., 1999, 274(8), HLA-DR alleles associated with rheumatoid arthritis and Proteus 5213-5221. mirabilis as the aetiological basis for autoimmunity. Microbes In- [120] Kato, R.; Kaga, C.; Kanie, K.; Kunimatsu, M.; Okochi, M.; Honda, fect., 2000, 2(12), 1489-1496. H. Peptide Array-Based Peptide-Cell Interaction Analysis. Mini- [102] Appel, H.; Mertz, A.; Distler, A.; Sieper, J.; Braun, J. The 19 kDa Rev Org. Chem., 2011, 8(2), 171-177. protein of Yersinia enterocolitica O:3 is recognized on the cellular [121] Kato, R.; Kaga, C.; Kunimatsu, M.; Kobayashi, T.; Honda, H. and humoral level by patients with Yersinia induced reactive arthri- Peptide arraybased interaction assay of solid-bound peptides and tis. J. Rheumatol., 1999, 26(9),1964-1971. anchorage-dependant cells and its effectiveness in cell-adhesive [103] Murphy, T.F.; Brauer, A.L. Expression of urease by Haemophilus peptide design. J. Biosci. Bioeng., 2006, 101(6), 485-495. influenzae during human respiratory tract infection and role in sur- [122] Falsey, J.R.; Renil, M.; Park, S.; Li, S.; Lam, K.S. Peptide and vival in an acid environment. BMC Microbiol., 2011, 11(1), 183. small molecule microarray for high throughput cell adhesion and [104] Contreras-Rodriguez, A.; Quiroz-Limon, Jose.; Martins, A.M.; functional assays. Bioconjug. Chem., 2001, 12(3), 346-353. Peralta, H.; Avila-Calderon, E.; Sriranganathan N.; Boyle, S.M.; [123] Thiele, A.; Pösel, S.; Spinka, M.; Zerweck, J.; Reimer, U.; Reineke, Lopez-Merino, A. Enzymatic, immunological and phylogenetic U.; Schutkowski, M. Profiling of Enzymatic Activities Using Pep- characterization of Brucella suis urease. BMC Microbiol., 2008, 8, tide Arrays. Mini-Reviews in Organic Chemistry, 2011, 8(2), 147- 121. 156. [105] Shin, D.-S.; Kim, D.-H.; Chung, W.-J.; Lee, Y.-S. Combinatorial [124] Leung, G.C.; Murphy, J.M.; Briant, D.; Sicheri, F. Characterization Solid Phase Peptide Synthesis and Bioassays. J. Biochem. Mol. of kinase target phosphorylation consensus motifs using peptide Biol., 2005, 38(5), 517-525. SPOT arrays. Methods Mol. Biol., 2009, 570, 187-195. [106] Benjamin, D.C.; Berzofsky, J.A.; East, I.J.; Gurd, F.R.; Hannum, [125] Houseman, B.T.; Huh, J.H.; Kron, S.J.; Mrksich, M. Peptide chips C.; Leach, S.J.; Margoliash, E.; Michael, J.G.; Miller, A.; Prager, for the quantitative evaluation of protein kinase activity. Nat. Bio- E.M.; Reichlin, M.; Sercarz, E.E.; Smith-Gill, S.J.; Todd, P.E.; technol., 2002, 20(3), 270-274. Wilson, A.C. The antigenic structure of proteins: a reappraisal. [126] Tegge, W.J.; Frank, R. Analysis of protein kinase substrate speci- Annu. Rev. Immunol., 1984, 2, 67-101. ficity by the use of peptide libraries on cellulose paper (SPOT- [107] Liu, R.; Enstrom, A.M.; Lam, K.S. Combinatorial peptide library method). Methods Mol. Biol., 1998, 87, 99-106. methods for immunobiology research. Exp. Hematol., 2003, 31(1), [127] Espanel, X.; Huguenin-Reggiani, M.; van Huijsduijnen, R.H. The 11-30. SPOT technique as a tool for studying protein tyrosine phosphatase [108] O’Brien-Simpson, N.M.; Pathirana, R.D.; Paolini, R.A.; Chen, Y- substrate specificities. Protein Sci., 2002, 11(10), 2326-2334. Y.; Veith, P.D.; Tam, V.; Ally, N.; Pike, R.N.; Reynolds, E.C. An [128] Moss, S.F.; Moise, L.; Lee, D.S.; Kim, W.; Zhang, S.; Lee, J; Immune Response Directed to Proteinase and Adhesin Functional Rogers, A.B.; Martin, W.; De Groot A.S. HelicoVax: epitope-based Epitopes Protects against Porphyromonas gingivalis-Induced therapeutic Helicobacter pylori vaccination in a mouse model. Periodontal Bone Loss. J. Immunol., 2005, 175(6), 3980-3989. Vaccine, 2011, 29(11), 2085-2091. [109] Frank, R. Spot-synthesis: an easy technique for the positionally [129] Hoffmann, S.; Frank, R. A new safety-catch peptide-resin linkage addressable, parallel chemical synthesis on a membrane support. for the direct release of peptides into aqueous buffers. Tetrahedron Tetrahedron, 1992, 48(42), 9217-9232. Lett., 1994, 35, 7763-7766;. [110] Mahler, M.; Fritzler, M.J. Epitope specificity and significance in [130] Panke, G.; Frank, R. Improved Preparation of a Safety-Catch systemic autoimmune diseases. Ann. N.Y. Acad. Sci., 2010, 1183, Linker for the Solid Phase Synthesis of Peptide Acids Finally Re- 267-287. leased into Aqueous Buffers. Tetrahedron Lett., 1998, 39(1), 17-18. [111] Schwemmle, M.; Billich, C. The use of peptide arrays for the char- [131] Scharn, D.; Wenschuh, H.; Reineke, U.; Schneider-Mergener, J.; acterization of monospecific antibody repertoires from polyclonal Germeroth, L. Spatially addressed synthesis of amino- and amino- sera of psychiatric patients suspected of infection by Borna disease oxy-substituted 1, 3,5-triazine arrays on polymeric membranes. J. virus. Mol. Divers., 2004, 8(3), 247-250. Comb. Chem., 2000, 2(4), 361-369. [112] Hilpert, K.; Hansen, G.; Wessner, H.; Küttner, G.; Welfle, K.; [132] Rau, H.K.; DeJonge, N.; Haehnel, W. Combinatorial Synthesis of Seifert, M.; Höhne, W. Anti-c-myc antibody 9E10: epitope key po- Four-Helix Bundle Hemoproteins for Tuning of Cofactor Proper- sitions and variability characterized using peptide spot synthesis on ties. Angew. Chem. Int. Ed. Engl., 2000, 39(1), 250-253. cellulose. Protein Eng., 2001, 14(10), 803-806. [133] Ay, B.; Landgraf, K.; Streitz, M.; Fuhrmann, S.; Volkmer, R.; [113] Tong, J.; Elowe, S.; Nash, P.; Pawson, T. Manipulation of EphB2 Boisguerin, P. Using hydroxymethylphenoxy derivates with the regulatory motifs and SH2 binding sites switches MAPK signaling SPOT technology to generate peptides with authentic C-termini. and biological activity. J. Biol. Chem., 2003, 278(8), 6111-6119. Bioorg. Med. Chem. Lett., 2008, 18(14), 4038-4043. [114] Yaffe, M.B.; Rittinger, K.; Volinia, S.; Caron, P.R.; Aitken, A.; [134] Kaca, W.; Kaminski, Z.J.; Kolesinska, B.; Kwinkowski, M.; Arab- Leffers, H.; Gamblin, S.J.; Smerdon, S.J.; Cantley, L.C. The struc- ski, M.; Konieczna, I. Peptides mimicking urease, methods of tural basis for 14-3-3: phosphopeptide binding specificity. Cell, manufacturing, application in diagnostic tests and the way of per- 1997, 91(7), 961-971. formance the test. Patent Applications PCT/PL2009/000106, [115] Nash, P.; Tang, X.; Orlicky, S.; Chen, Q.; Gertler, F.B.; Menden- WO/2010/071462, December 16, 2009. hall, M.D.; Sicheri, F.; Pawson, T.; Tyers, M. Multisite phosphory- [135] Kami ski, Z.J.; Paneth, P.; Rudzi ski, J. A study on the activation lation of a CDK inhibitor sets a threshold for the onset of DNA rep- of carboxylic acids by means of 2-chloro-4,6-dimethoxy-1,3,5- lication. Nature, 2001, 414(6863), 514-521. triazine and 2-chloro-4,6-diphenoxy-1,3,5-triazine. J. Org. Chem., [116] Espanel, X.; Wälchli, S.; Rückle, T.; Harrenga, A.; Huguenin- 1998, 63, 4248-4255. Reggiani, M.; van Huijsduijnen, R.H. Mapping of synergistic com- [136] Kami ski, Z.J.; Paneth, P.; O'Leary, M. Nitrogen Isotope Effects on ponents of weakly interacting protein-protein motifs using arrays of the Acylation of Aniline J. Org. Chem., 1991, 56, 5716-5719. paired peptides. J. Biol. Chem., 2003, 278(17), 15162-15167. [137] Kami ski, Z.J.; G ówka, M.L.; Olczak, A.; Martynowski, D. Ther- [117] Huang, H.; Li, L.; Wu, C.; Schibli, D.; Colwill, K.; Ma, S.; Li, C.; mal isomerization of 2- acyloxy-4,6-dimethoxy-1,3,5-triazines to 1- Roy, P.; Ho, K.; Songyang, Z.; Pawson, T.; Gao, Y.; Li, S.S.-C. acyl-3,5-dimethyl-1,3,5-triazin-2,4,6(1H,3H,5H)-triones. Crystal Defining the specificity space of the human SRC homology 2 do- structure of 1-(2,2-dimethylpropanoyloxy)3,5-dimethyl-1,3,5- main. Mol. Cell. Proteomics, 2008, 7(4), 768-784. triazin-2,4,6(1H,3H,5H)-trione. Pol. J. Chem., 1996, 70, 1316- [118] Smith, M.J.; Hardy, W.R.; Murphy, J.M.; Jones, N.; Pawson, T. 1323. Screening for PTB domain binding partners and ligand specificity [138] Kolesi ska, B.; Grabowski, S.; Konieczna, I.; Kaca, W.; Peroni, E.; using proteomederived NPXY peptide arrays. Mol. Cell Biol., Papini, A.M.; Rovero, P.; Kami ski, Z.J. In: Peptides 2006, Pro- 2006, 26(22), 8461-8474. ceedings of the 29th European Peptide Symposium, Poland, Sep- [119] Reuter, M.; Schneider-Mergener, J.; Kupper, D.; Meisel, A.; Mack- tember 3-8, 2006; Rolka, K.; Rekowski, P.; Silberring, J.; Eds., eldanz, P.; Krüger, D.H.; Schroeder, C. Regions of endonuclease Kenes International: Geneva, 2007, 616-617. EcoRII involved in DNA target recognition identified by mem- 806 Current Protein and Peptide Science, 2012, Vol. 13, No. 8 Konieczna et al. [139] Glenska, J; Adamus-Bia ek W., Kwinkowski M., Kolesi ska B., ter pylori bound by neutralizing antibodies. Vaccine, 2010, 28, Kami ski Z., Kaca W. In: Abstracts of the VI Congress of Polish 5220-5227. Biotechnology “IV EUROBIOTECH 2011” and Central European [142] Flach, C.F.; Svensson, N.; Blomquist, M.; Ekman, A.; Raghavan, Congress of Live Science, Kraków, Poland, October 12-15, 2011; S.; Holmgren, J. A truncated form of HpaA is a promising antigen Acta Biochim. Pol., 2011; 58, Supp. 4, 67. for use in a vaccine against Helicobacter pylori. Vaccine, 2011, [140] Fujii, R.; Morihara, F.; Oku, T.; Hifumi, E.; Uda, T. Epitope map- 29(6), 1235-1241. ping and features of the epitope for monoclonal antibodies inhibit- [143] Guo, L.; Li, X.; Tang, F.; He, Y.; Xing, Y.; Deng, X.; Xi, T. Im- ing enzymatic activity of Helicobacter pylori urease. Biotechnol. munological features and the ability of inhibitory effects on enzy- Bioeng., 2004, 86(4), 434-44. matic activity of an epitope vaccine composed of cholera toxin B [141] Qiua, Y.; Wanga, Y-C.; Taoa, H-X.; Zhana, D-W.; Yuana, S-L.; subunit and B cell epitope from Helicobacter pylori urease A Wanga, P.; Wanga, L-C.; Hanb, X-P.; Li, C-S.; Li, J-K.; Liua, C-J. subunit. Appl. Microbiol. Biotechnol., 2012, 93(5), 1937-1945. Identification of B-cell epitopes in urease B subunit of Helicobac- Received: June 22, 2012 Revised: July 15, 2012 Accepted: September 03, 2012

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