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Identification and Biochemical Characterization of a New Xylan-degrading Streptomyces atrovirens Subspecies WJ-2 Isolated from Soil of Jeju Island in Korea

제주도 토양으로부터 자일란 분해 Streptomyces atrovirens subspecies WJ-2 동정 및 효소의 생화학적 특성 규명

  • Kim, Da Som (Biological and Genetic Resources Assessment Division, National Institute of Biological Resources) ;
  • Bae, Chang Hwan (Biological and Genetic Resources Assessment Division, National Institute of Biological Resources) ;
  • Yeo, Joo Hong (Biological and Genetic Resources Assessment Division, National Institute of Biological Resources) ;
  • Chi, Won-Jae (Biological and Genetic Resources Assessment Division, National Institute of Biological Resources)
  • 김다솜 (국립생물자원관 생물자원활용부 유용자원분석과) ;
  • 배창환 (국립생물자원관 생물자원활용부 유용자원분석과) ;
  • 여주홍 (국립생물자원관 생물자원활용부 유용자원분석과) ;
  • 지원재 (국립생물자원관 생물자원활용부 유용자원분석과)
  • Received : 2016.09.12
  • Accepted : 2016.10.17
  • Published : 2016.12.28

Abstract

A bacterial strain was isolated from a soil sample collected on Jeju Island, Korea. The strain, designated WJ-2, exhibited a high xylanase activity, whereas cellulase activity was not detected. The 16S rRNA gene sequence of WJ-2 was highly similar to type strains of the genus Streptomyces. A neighbor-joining phylogenetic tree based on 16S rRNA gene sequences showed that strain WJ-2 is phylogenetically related to Streptomyces atrovirens. Furthermore, DNA-DNA hybridization analysis confirmed that strain WJ-2 is a novel subspecies of Streptomyces atrovirens. The genomic DNA G+C content was 73.98 mol% and the major fatty acid present was anteiso-C15:0 (36.19%). The growth and xylanase production of strain WJ-2 were significantly enhanced by using soytone and xylan as nitrogen and carbon sources, respectively. Crude enzyme preparations from the culture broth of strain WJ-2 exhibited maximal total xylanase activities at pH 7.0 and $55^{\circ}C$. Thin-layer chromatography analysis revealed that the crude enzyme degrades beechwood xylan to yield xylobiose and xylotriose as the principal hydrolyzed end products.

제주도에서 채집된 토양시료로부터 xylanase 활성을 나타내는 균주를 분리하여 WJ-2로 명명하였다. 균주 WJ-2의 16S rRNA 유전자 염기서열을 결정하여 이를 토대로 상동성을 검색한 결과, Streptomyces 속의 균주들과 높은 염기서열 상동성을 보였다. 16S rRNA 유전자 염기서열을 토대로하는 neighbor-joining 계통수를 제작하여 Streptomyces atrovirens와 가장 높은 계통발생적 연관성이 갖고 있는 것을 밝혔다. 또한 DNA-DNA hybridization 분석을 통하여 Streptomyces atrovirens의 신규한 아종임을 증명하였다. 균주 WJ-의 게놈내 GC 농도는 73.98 mol%이었으며, 주요 세포벽 지방산으로 anteiso-$C_{15:0}$ (36.19%)을 함유하고 있었다. 균주 WJ-2의 성장 및 xylanase 생산은 배지내에 질소원으로 soytone과 탄소원으로 xylan을 첨가하였을 때 급격히 증가되는 것을 확인하였다. 액체배양액으로부터 준비된 조효소의 xylanase 활성은 pH 7.0과 $55^{\circ}C$에서 가장 높게 나타났다. Thin layer chromatography (TLC) 분석을 통하여 균주 WJ-2의 조효소는 xylan을 분해하여 최종분해산물로서 xylobiose와 xylotriose 생산하는 효소임을 확인하였다.

Keywords

References

  1. Achary AA, Prapulla SG. 2009. Value addition to corncob: Production and characterization of xylo-oligosaccharides from alkali pretreated lignin-saccharide complex using Aspergillus oryzae MTCC5154. Bioresour. Technol. 100: 991-995. https://doi.org/10.1016/j.biortech.2008.06.050
  2. Al-Bari MAA, Bhuiyan MSA, Flores ME, Petrosyan P, Garcia-Varela M, Islam MAU. 2005. Streptomyces bangladeshensis sp. nov., isolated from soil, which produces bis-(2-ethylhexyl) phthalate. Int. J. Syst. Evol. Microbiol. 55: 1973-1977. https://doi.org/10.1099/ijs.0.63516-0
  3. Altschul SF, Madden TL, Schaffer AA, Zhang J, Zhang A, Miller W, et al. 1990. Gapped BLAST and PSI-BLAST: a new generation of protein database search programs. Nucleic Acids Res. 25: 3389-402.
  4. Bajaj BK, Singh NP. 2010. Production of xylanase from an alkalitolerant Streptomyces sp. 7b under solid-state fermentation, its purification, and characterization. Appl. Biochem. Biotechnol. 162: 1804-1818. https://doi.org/10.1007/s12010-010-8960-x
  5. Baker GC, Smith JJ, Cowan DA. 2003. Review and re-analysis of domain-specific 16S primers. J. Microbiol. Methods 55: 541-555. https://doi.org/10.1016/j.mimet.2003.08.009
  6. Beg QK, Kapoor M, Mahajan L, Hoondal GS. 2001. Microbial xylanases and their industrial applications: a review. Appl. Microbiol. Biotechnol. 56: 326-338. https://doi.org/10.1007/s002530100704
  7. Biely P. 1985. Microbial xylanolytic systems. Trends Biotechnol. 11: 286-290.
  8. Brennan Y, Callen WN, Christoffersen L, Dupree P, Goubet F, Healey S, et al. 2004. Unusual microbial xylanases from insect guts. Appl. Environ. Microbiol. 70: 3609-3617. https://doi.org/10.1128/AEM.70.6.3609-3617.2004
  9. Chi WJ, Lim JH, Park DY, Park JS, Hong SK. 2013. Production and characterization of a thermostable endo-type $\beta$-xylanase produced by a newly-isolated Streptomyces thermocarboxydus subspecies MW8 strain from Jeju Island. Proc. Biochem. 48: 1736-1743. https://doi.org/10.1016/j.procbio.2013.07.024
  10. Chun J, Lee JH, Jung YY, Kim MJ, Kim SI, Kim BK, et al. 2007. EzTaxon: a web-based tool for the identification of prokaryotes based on 16S ribosomal RNA gene sequences. Int. J. Syst. Evol. Microbiol. 57: 2259-2261. https://doi.org/10.1099/ijs.0.64915-0
  11. Dhiman SS, Sharma J, Battan B. 2008. Industrial applications and future prospects of microbial xylanases: a review. Bioresources 3: 1377-1402.
  12. Elegir G, Szakacs G, Jeffries TW. 1994. Purification, characterization, and substrate specificities of multiple xylanases from Streptomyces sp. Strain B-12-2. Appl. Environ. Microbiol. 60: 2609-2615.
  13. El-Sersy NA, Abd-Elnaby H, Abou-Elela GH, Ibrahim AHA, El-Toukhy NMK. 2010. Optimization, economization and characterization of cellulase produced by marine Streptomyces ruber. Afr. J. Biotechnol. 9: 6355-6364.
  14. Gause GF, Preobrazhenskaya TP, Sveshnikova MA, Terekhova LP, Maximova TS. 1983. A guide for the determination of actinomycetes. Genera Streptomyces, Streptoverticillium, and Chainia. Moscow, USSR: Nauka.
  15. Georis J, Giannotta F, Buyl ED, Granier B, Frere JM. 2000. Purification and properties of three endo-$\beta$-1,4-xylanases produced by Streptomyces sp. strain S38 which differ in their ability to enhance the bleaching of kraft pulps. Enzyme Microb. Technol. 26: 178-186. https://doi.org/10.1016/S0141-0229(99)00141-6
  16. Grabski AC, Forrester IT, Patel R, Jeffries TW. 1993. Characterization and N-terminal amino acid sequences of beta-(1-4) endoxylanases from Streptomyces roseiscleroticus: purification incorporating a bioprocessing agent. Protein Expr. Purif. 4: 120-129. https://doi.org/10.1006/prep.1993.1018
  17. Gupta S, Kuhad RC, Bhushan B, Hoondal GS. 2001. Improved xylanase production from a haloalkaliphilic Staphylococcus sp. SG-13 using inexpensive agricultural residues. World J. Microbiol. Biotechnol. 17: 5-8. https://doi.org/10.1023/A:1016691205518
  18. Hopwood DA, Bibb MJ, Chater KF, Kieser T, Bruton CJ, Lydiate DJ, et al. 1985. Genetic manipulation of Streptomyces, A Laboratory Manual, Norwich, UK: The John Innes Foundation.
  19. Hwang IT, Lim HK, Song HY, Cho SJ, Chang JS, Park NJ. 2010. Cloning and characterization of a xylanase, KRICT PX1 from the strain Paenibacillus sp. HPL-001. Biotechnol. Adv. 28: 594-601. https://doi.org/10.1016/j.biotechadv.2010.05.007
  20. Kallel F, Driss D, Chaabouni SE, Ghorbel R. 2015. Biological activities of xylooligosaccharides generated from garlic straw xylan by purified xylanase from Bacillus mojavensis UEB-FK. Appl. Biochem. Biotechnol. 175: 950-964. https://doi.org/10.1007/s12010-014-1308-1
  21. Kieser H, Bibb MJ, Buttner MJ, Chater KF, Hopwood DA. 2000. Practical Streptomyces genetics. The John Innes Foundation, Norwich, United Kingdom.
  22. Kimura M. 1983. The neutral theory of molecular evolution. Cambridge, UK: Cambridge University Press.
  23. La Duc MT, Kern R, Venkateswaran K. 2004. Microbial monitoring of spacecraft and associated environments. Microbial. Ecol. 47: 150-158. https://doi.org/10.1007/s00248-003-1012-0
  24. Lee CC, Kibblewhite-Accinelli RE, Wagschal K, Robertson GH, Wong DW. 2006. Cloning and characterization of a cold-active xylanase enzyme from an environmental DNA library. Extremophiles 10: 295-300. https://doi.org/10.1007/s00792-005-0499-3
  25. Mesbah M, Premachandran U, Whitman WB. 1989. Precise measurement of the G+C content of deoxyribonucleic acid by high-performance liquid chromatography. Int. J. Syst. Bacteriol. 39: 159-167. https://doi.org/10.1099/00207713-39-2-159
  26. Miller GL. 1959. Use of dinitrosalicylic acid reagent for determination of reducing sugar. Anal. Chem. 31: 426-428. https://doi.org/10.1021/ac60147a030
  27. Nath D, Rao M. 2001. pH dependent conformational and structural changes of xylanase from an alkalophilic thermophilic Bacillus sp (NCIM 59). Enzyme Microb. Technol. 28: 397-403. https://doi.org/10.1016/S0141-0229(00)00359-8
  28. Ninawe S, Kapoor M, Kuhad RC. 2008. Purification and characterization of extracellular xylanase from Streptomyces cyaneus SN32. Bioresour. Technol. 99: 1252-1258. https://doi.org/10.1016/j.biortech.2007.02.016
  29. Ruiz-Arribas A, Fernandez-Abalos JM, Sanchez P, Garda AL, Santamaria RI. 1995. Overproduction, purification, and biochemical characterization of a xylanase (Xys1) from Streptomyces halstedii JM8. Appl. Environ. Microbiol. 35: 2414-2419.
  30. Satomi M, Kimura B, Hamada T, Harayama S, Fujii T. 2002. Phylogenetic study of the genus Oceanospirillum based on 16S rRNA and gyrB genes: emended description of the genus Oceanospirillum, description of Pseudospirillum gen. nov., Oceanobacter gen. nov. and Terasakiella gen. nov. and transfer of Oceanospirillum jannaschii and Pseudomonas stanieri to Marinobacterium as Marinobacterium jannaschii comb. nov. and Marinobacterium stanieri comb. nov. Int. J. Syst. Evol. Microbiol. 52: 739-747.
  31. Shin JH, Choi JH, Lee OS, Kim YM, Lee DS, Kwak YY, et al. 2009. Thermostable xyalanse from Streptomyces thermocyaneoviolaceus for optimal production of xylooligosaccharides. Biotechnol. Bioprocess Eng. 14: 391-399. https://doi.org/10.1007/s12257-008-0220-3
  32. Subramanian S, Sandhia GS, Prema P. 2001. Control of xylanase production without protease activity in Bacillus sp. by selection of nitrogen source. Biotechnol. Lett. 23: 369-371. https://doi.org/10.1023/A:1005663704321
  33. Techapum C, Charoenrat T, Watanabe M, Sasaki K, Poosara N. 2002. Optimization of thermostable and alkaline-tolerant cellulase-free xylanase production from agricultural waste by thermotolerant Streptomyces sp. Ab106, using the central composite experimental design. Biochem. Eng. J. 12: 99-105. https://doi.org/10.1016/S1369-703X(02)00047-5
  34. Temuujin U, Chi WJ, Lee SY, Chang YK, Hong SK. 2011. Overexpression and biochemical characterization of DagA from Streptomyces coelicolor A3(2): an endo-type $\beta$-agarase producing neoagarotetraose and neoagarohexaose. Appl. Microbiol. Biotechnol. 92: 749-759. https://doi.org/10.1007/s00253-011-3347-7
  35. Van Trappen S, Tan TL, Yang J, Mergaert J, Swings J. 2004. Altermonas stellipolaris sp. nov., a novel, budding, prosthecate bacterium from Antarctic seas, and emended description of the genus Altermonas. Int. J. Syst. Evol. Microbiol. 54: 1157-1163. https://doi.org/10.1099/ijs.0.02862-0
  36. Vazquez MJ, Alonso JL, Dominguez H, Parajo JC. 2001. Xylooligosaccharides: Manufacture and applications. Trends Food Sci. Technol. 11: 387-393.
  37. Wang SL, Yen YH, Shih IL, Chang AC, Chang WT, Wu WC, et al. 2003. Production of xylanases from rice bran by Streptomyces actuosus A-151. Enzyme Microb. Technol. 33: 917-925. https://doi.org/10.1016/S0141-0229(03)00246-1
  38. Yan Q, Hao S, Jiang Z, Zhai Q, Chen W. 2009. Properties of a xylanase from Streptomyces matensis being suitable for xylooligosaccharides production. J. Mol. Catal. B Enzym. 58: 72-77. https://doi.org/10.1016/j.molcatb.2008.11.010