Access the full text.
Sign up today, get DeepDyve free for 14 days.
Ann Microbiol (2017) 67:391–393 DOI 10.1007/s13213-017-1267-1 SHORT COMMUNICATION Genomic analysis of type strain Paenibacillus aceti L14 , a highly efficient producer of pyrazines 1,2 2 2 1 Pan Li & Xing Gan & Lixin Luo & Bing Du Received: 20 December 2016 /Accepted: 19 April 2017 /Published online: 28 April 2017 Springer-Verlag Berlin Heidelberg and the University of Milan 2017 Abstract Paenibacillus aceti L14 (CGMCC 1.15420 = JCM Findings 31170) is a novel species isolated from the solid-state acetic acid fermentation culture of traditional Chinese vinegar. The Pyrazines occur ubiquitously in nature and can be strain is able to biosynthesize the pyrazines, including 2,3- biosynthesized by several bacteria. Pyrazines have paramount diisobutylpyrazine, 2-isobutyl-3-methylpyrazine and 1-(5- biological activities and are important flavour additives in the isobutyl-2-pyrazinyl)-1-propanone. Genome sequencing of food industry. However, knowledge about the biosynthesis of L14 was performed to gain insights into the genetic elements pyrazines is very limited. Nowadays, two different pathways involved in the biosynthesis of pyrazines. The genome of L14 for the biosynthesis of alkyl-substituted pyrazines have been contains 5,611,962 bp with a GC content of 47.92 mol%, proposed (Beck et al. 2003): (i) the pyrazine ring structure is 5147 protein coding genes, 92 tRNAs, 20 rRNAs and four formed by the condensation of the amidated amino acid with sRNAs. The strain L14 also contains complete biosynthetic an α,β-dicarbonyl compound and, followed by methylation pathways of valine, leucine and isoleucine, and contains genes reaction, leads to alkyl- and methoxy-substituted pyrazines for encoding threonine dehydratase and ketol-acid (Murray and Whitfield 1975); (ii) pyrazine biosynthesis is reductoisomerase. This genome sequence provides a basis the condensation of two amino acids to a cyclic dipeptide, for elucidating the possible mechanism for the biosynthesis and then converted into the pyrazine molecule (Cheng et al. of pyrazines. 1991). The genus Paenibacillus was firstly proposed as a new group of bacilli by Ash et al. (1993) based on the analysis of . . . Keywords Paenibacillus aceti Genome Pyrazine 16S rRNA gene sequences. Members of this genus are gener- Leucine Threonine dehydratase ally Gram-positive, rod-shaped, endospore-forming, aerobic or facultatively anaerobic bacteria. Nowadays, numerous members of the genus Paenibacillus have been isolated from various environments with very diverse biochemical functions (Zhang et al. 2013; Xie et al. 2014). In our previous study, we isolated a novel species of the genus Paenibacillus from the traditional solid-state acetic acid * Lixin Luo fermentation culture of Chinese cereal vinegar and designated email@example.com as Paenibacillus aceti L14 (Li et al. 2016a). To further char- * Bing Du acterise several features of strain L14, the strain P. aceti L14 firstname.lastname@example.org was initially pre-inoculated with 5 mL tryptic soy broth (TSB; Difco) at 37 °C for 24 h to obtain seed culture. Afterwards, College of Food Science, South China Agricultural University, 1 mL of seed culture was inoculated in 100 mL tryptic soy Wushan Road, Tianhe District, Guangzhou 510642, China broth (TSB; Difco) and the flasks were shaken at 37 °C and Guangdong Key Laboratory of Fermentation and Enzyme 200 rpm for 24 h. The control culture was designed as follows: Engineering, School of Bioscience and Bioengineering, South China University of Technology, Guangzhou 510006, China 1 mL tryptic soy broth was inoculated in 100 mL tryptic soy 392 Ann Microbiol (2017) 67:391–393 Table 1 Genome broth (TSB; Difco) and was also treated under the same con- Feature Value features of Paenibacillus ditions. The supernatant culture was obtained by centrifuging aceti L14 at 8000×g and 4 °C for 5 min. The aroma compound in the Genome size (bp) 5,611,962 supernatant was detected by headspace solid-phase GC content 47.92% microextraction (HS-SPME) coupled with gas chromatogra- Gene total length (bp) 4,731,522 phy–mass spectrometry (GC-MS) (Li et al. 2016b). Number of scaffolds 11 According to the results of HS-SPME-GC-MS, this strain is Number of contigs 64 able to biosynthesize the pyrazines (1.68 g/L), including 2,3- Largest scaffold (bp) 5,572,757 diisobutylpyrazine, 2-isobutyl-3-methylpyrazine and 1-(5- Largest contig (bp) 773,790 isobutyl-2-pyrazinyl)-1-propanone (Fig. 1). Protein-coding genes 5147 In order to gain insights into the genetic bases in the bio- tRNA genes 92 synthesis of pyrazines, the genome of P. aceti L14 was se- rRNA genes 20 quenced on an Illumina HiSeq 2500 sequencing platform. A sRNA genes 4 500-bp paired-end library and a 6-kbp mate-pair library were Minisatellite DNA 100 constructed using the TruSeq DNA PCR-Free Sample Genes with predicted 4789 Preparation Kit (Illumina) and the Nextera Mate Pair Sample function Preparation Kit (Illumina), respectively. This strategy pro- duced 2053 Mb of paired-end data with about 339-fold cov- erage of the genome. After data processing and assembly by et al. 2007) and the Rfam database (Gardner et al. 2009). In total, 5147 protein coding genes, 92 tRNAs, 20 rRNAs and applying SOAPdenovo 2.04 (Luo et al. 2012), the P. aceti L14 chromosome consisted of 11 scaffolds comprising 64 four sRNAs were determined (Table 1). scaffolded contigs. The genome has a size of 5,611,962 bp, Ammonium was found to be the precursor for the biosyn- featuring a GC content of 47.92% (Table 1). The coding se- thesis of alkylated pyrazine (Zhu and Xu 2010). We annotated quences were predicted by GeneMarkS (Besemer et al. 2001) 50 CDSs for peptidase and four CDSs for amino acid dehy- drogenase, which can degrade proteins and peptides into ami- and annotated using public databases, including the non- redundant (NR), KEGG, COG, GO and Swiss-Prot databases. no acids and produce ammonium from amino acids, respec- tively. Valine, leucine and isoleucine were the precursors of The tRNA, rRNA and sRNA genes were identified by tRNAscan-SE (Lowe and Eddy 1997), RNAmmer (Lagesen the isobutyl- or isopropyl-substituted pyrazines (Beck et al. Fig. 1 Gas chromatography–mass spectrometry (GC-MS) chromatograms of fermentation culture (a)of Paenibacillus aceti L14 and control culture (b) for volatile compounds obtained by headspace solid-phase microextraction (HS-SPME). 2-Octanol was used as the internal standard Ann Microbiol (2017) 67:391–393 393 Beck HC, Hansen AM, Lauritsen FR (2003) Novel pyrazine metabolites 2003). Notably, the complete biosynthetic pathways were an- found in polymyxin biosynthesis by Paenibacillus polymyxa. FEMS notated for the production of valine, leucine and isoleucine in Microbiol Lett 220:67–73 strain L14. The genes predicted for encoding threonine Besemer J, Lomsadze A, Borodovsky M (2001) GeneMarkS: a self- dehydratase and ketol-acid reductoisomerase, which are re- training method for prediction of gene starts in microbial genomes. Implications for finding sequence motifs in regulatory regions. quired for the formation of 2-oxobutanoate and (S)-2- Nucleic Acids Res 29:2607–2618 acetolactate, have proven to be indispensable for the biosyn- Cheng TB, Reineccius GA, Bjorklund JA, Leete E (1991) Biosynthesis of thesis of pyrazines through the valine and leucine biosynthetic 2-methoxy-3-isopropylpyrazine in pseudomonas perolens. J Agric pathway in Corynebacterium glutamicum (Dickschat et al. Food Chem 39:1009–1012 Dickschat JS, Wickel S, Bolten CJ, Nawrath T, Schulz S, Wittmann C 2010). (2010) Pyrazine biosynthesis in Corynebacterium glutamicum.Eur In conclusion, the genome data of P. aceti L14 is helpful for J Org Chem 2010:2687–2695 uncovering the possible mechanism for the biosynthesis of Gardner PP, Daub J, Tate JG, Nawrocki EP, Kolbe DL, Lindgreen S, pyrazines, and will facilitate its potential applications as starter Wilkinson AC, Finn RD, Griffiths-Jones S, Eddy SR, Bateman A (2009) Rfam: updates to the RNA families database. Nucleic Acids cultures in the food industry. Res 37:D136–D140 Lagesen K, Hallin P, Rødland EA, Stærfeldt H-H, Rognes T, Ussery DW (2007) RNAmmer: consistent and rapid annotation of ribosomal RNA genes. Nucleic Acids Res 35:3100–3108 Strain and nucleotide sequence accession numbers Li P, Lin W, Liu X, Li S, Luo L, Lin W-T (2016a) Paenibacillus aceti sp. nov., isolated from the traditional solid-state acetic acid fermentation This whole genome shotgun project has been deposited at culture of Chinese cereal vinegar. Int J Syst Evol Microbiol 66: DDBJ/ENA/GenBank under the accession number 3426–3431 Li S, Li P, Liu X, Luo L, Lin W (2016b) Bacterial dynamics and metab- MDDO00000000. The BioProject designation for this project olite changes in solid-state acetic acid fermentation of Shanxi aged is PRJNA338379. The strain is available at the China General vinegar. Appl Microbiol Biotechnol 100:4395–4411 Microbiological Culture Collection Center and the Japan Lowe TM, Eddy SR (1997) tRNAscan-SE: a program for improved de- Collection of Microorganisms under the accession numbers tection of transfer RNA genes in genomic sequence. Nucleic Acids CGMCC 1.15420 and JCM 31170, respectively. Res 25:955–964 Luo R, Liu B, Xie Y, Li Z, Huang W, Yuan J, He G, Chen Y, Pan Q, Liu Y, Tang J, Wu G, Zhang H, Shi Y, Liu Y, Yu C, Wang B, Lu Y, Han C, Acknowledgements This research was supported by the National Cheung DW, Yiu S-M, Peng S, Xiaoqian Z, Liu G, Liao X, Li Y, Natural Science Foundation of China (grant no. 31271924). Yang H, Wang J, Lam T-W, Wang J (2012) SOAPdenovo2: an empirically improved memory-efficient short-read de novo assem- bler. GigaScience 1:18 Compliance with ethical standards Murray KE, Whitfield FB (1975) The occurrence of 3-alkyl-2- methoxypyrazines in raw vegetables. J Sci Food Agric 26:973–986 Conflict of interest The authors declare that they have no conflict of XieJ-B,DuZ,Bai L, Tian C, ZhangY,Xie J-Y, Wang T, LiuX,ChenX, interest. Cheng Q, Chen S (2014) Comparative genomic analysis of N 2-fixing and non-N 2-fixing Paenibacillus spp.: organization, evolution and expression of the nitrogen fixation genes. PLoS Genet 10:e1004231 Zhang J, Wang Z-T, Yu H-M, Ma Y (2013) Paenibacillus catalpae sp. References nov., isolated from the rhizosphere soil of Catalpa speciosa.IntJ Syst Evol Microbiol 63:1776–1781 Ash C, Priest FG, Collins MD (1993) Molecular identification of rRNA Zhu B-F, Xu Y (2010) A feeding strategy for tetramethylpyrazine pro- group 3 bacilli (ash, farrow, Wallbanks and Collins) using a PCR duction by Bacillus subtilis based on the stimulating effect of am- probe test. Anton Leeuw Int J G 64:253–260 monium phosphate. Bioprocess Biosyst Eng 33:953–959
Annals of Microbiology – Springer Journals
Published: Apr 28, 2017
Access the full text.
Sign up today, get DeepDyve free for 14 days.