Gene cloning, expression and characterization of a new cold-active and salt-tolerant endo-β-1,4-xylanase from marine Glaciecola mesophila KMM 241

  • Bing Guo
  • Xiu-Lan Chen
  • Cai-Yun Sun
  • Bai-Cheng Zhou
  • Yu-Zhong Zhang
Biotechnologically Relevant Enzymes and Proteins

Abstract

Although a lot of xylanases are studied, only a few xylanases from marine microorganisms have been reported. A new xylanase gene, xynA, was cloned from marine bacterium Glaciecola mesophila KMM 241. Gene xynA contains 1,272 bp and encodes a 423-amino acid xylanase precursor. The recombinant xylanase, XynA, expressed in Escherichia coli BL21 is a monomer with a molecular mass of 43 kDa. Among the characterized xylanases, XynA shares the highest identity (46%) to the xylanase from Flavobacterium sp. strain MSY2. The optimum pH and temperature for XynA is 7.0 and 30 °C. XynA retains 23% activity and 27% catalytic efficiency at 4 °C. XynA has low thermostability, remaining 20% activity after 60-min incubation at 30 °C. Its apparent melting temperature (T m) is 44.5 °C. These results indicate that XynA is a cold-active xylanase. XynA shows a high level of salt-tolerance, with the highest activity at 0.5 M NaCl and retaining 90% activity in 2.5 M NaCl. It may be the first salt-tolerant xylanase reported. XynA is a strict endo-β-1,4-xylanase with a demand of at least four sugar moieties for effective cleavage. It efficiently hydrolyzes xylo-oligosaccharides and xylan into xylobiose and xylotriose without producing xylose, suggesting its potential in xylo-oligosaccharides production.

Keywords

Xylanase XynA Cold-active Salt-tolerant Glaciecola mesophila Marine Xylan 

Notes

Acknowledgment

The work was supported by Hi-Tech Research and Development program of China (2007AA091504, 2007AA091903), COMRA Program (DYXM-115-02-2-6), and Specialized Research Fund for the Doctoral Program of Higher Education (20060422053).

Supplementary material

253_2009_2056_MOESM1_ESM.doc (348 kb)
ESM 1 (DOC 347 kb)

References

  1. Akila G, Chandra TS (2003) A novel cold-tolerant Clostridium strain PXYL1 isolated from a psychrophilic cattle manure digester that secretes thermolabile xylanase and cellulase. FEMS Microbiol Lett 219:63–67CrossRefGoogle Scholar
  2. Ali MK, Rudolph FB, Bennett GN (2004) Thermostable xylanase 10B from Clostridium acetobutylicum ATCC824. J Ind Microbiol Biotechnol 31:229–234CrossRefGoogle Scholar
  3. Araki T, Tani S, Maeda K, Hashikawa S, Nakagawa H, Morishita T (1999) Purification and characterization of β-1, 3-xylanase from a marine bacterium, Vibrio sp. XY-214. Biosci Biotechnol Biochem 63:2017–2019CrossRefGoogle Scholar
  4. Badal CS (2002) Production, purification and properties of xylanase from a newly isolated Fusarium proliferatum. Process Biochem 37:1279–1284CrossRefGoogle Scholar
  5. Biely P, Vrsanská M, Tenkanen M, Kluepfel D (1997) Endo-β-1, 4-xylanase families: differences in catalytic properties. J Biotechnol 57:151–166CrossRefGoogle Scholar
  6. Blanco J, Coque JJR, Velasco J, Martín JF (1997) Cloning, expression in Streptomyces lividans and biochemical characterization of a thermostable endo-β-1, 4-xylanase of Thermomonospora alba ULJB1 with cellulose-binding ability. Appl Microbiol Biotechnol 48:208–217CrossRefGoogle Scholar
  7. Bradford MM (1976) A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem 72:248–254CrossRefGoogle Scholar
  8. Bradner JR, Sidhu RK, Gillings M, Nevalainen KM (1999) Hemicellulase activity of antarctic microfungi. J Appl Microbiol 87:366–370CrossRefGoogle Scholar
  9. Collins T, Meuwis MA, Stals I, Claeyssens M, Feller G, Gerday C (2002) A novel family 8 xylanase: functional and physico-chemical characterization. J Biol Chem 277:35133–35139CrossRefGoogle Scholar
  10. Collins T, Gerday C, Feller G (2005) Xylanases, xylanase families and extremophilic xylanases. FEMS Microbiol Rev 29:3–23CrossRefGoogle Scholar
  11. Collins T, Hoyoux A, Dutron A, Georis J, Genot B, Dauvrin T, Arnaut F, Gerday C, Feller G (2006) Use of glycoside hydrolase family 8 xylanases in baking. J Cereal Sci 43:79–84CrossRefGoogle Scholar
  12. Fialho MB, Carmona EC (2004) Purification and characterization of xylanase from Aspergillus giganteus. Folia Microbiol 49:13–18CrossRefGoogle Scholar
  13. Gilkes NR, Henrissat B, Kilburn DG, Miller RC Jr, Warren RA (1991) Domains in microbial 4-glycanases: sequence conservation, function, and enzyme families. Microbiol Rev 55:303–315Google Scholar
  14. Gruppen H, Hamer RJ, Voragen AGJ (1992) Water-unextractable cell wall material from wheat flour. 2. Fractionation of alkali-extracted polymers and comparison with waterextractable arabinoxylans. J Cereal Sci 16:53–67CrossRefGoogle Scholar
  15. Gupta S, Bhushan B, Hoondal GS (2000) Isolation, purification and characterization of xylanase from Staphylococcus sp. SG-13 and its application in biobleaching of kraft pulp. J Appl Microbiol 88:325–334CrossRefGoogle Scholar
  16. Henrissat B (1991) A classification of glycosyl hydrolases based on amino acid sequence similarities. Biochem J 280:309–316Google Scholar
  17. Hou YH, Wang TH, Long H, Zhu HY (2006) Novel cold-adaptive Penicillium strain FS010 secreting thermo-labile xylanase isolated from Yellow Sea. Acta Biochim Biophys Sin 38(2):142–149CrossRefGoogle Scholar
  18. Hu Y, Zhang G, Li A, Chen J, Ma L (2008) Cloning and enzymatic characterization of a xylanase gene from a soil-derived metagenomic library with an efficient approach. Appl Microbiol Biotechnol 80:823–830CrossRefGoogle Scholar
  19. Humphry DR, George A, Black GW, Cummings SP (2001) Flavobacterium frigidarium sp. nov., an aerobic, psychrophilic, xylanolytic and laminarinolytic bacterium from Antarctica. Int J Syst Evol Microbiol 51:1235–1243Google Scholar
  20. Inglis GD, Popp AP, Selinger LB, Kawchuk LM, Gaudet DA, McAllister TA (2000) Production of cellulases and xylanases by low-temperature basidiomycetes. Can J Microbiol 46:860–865CrossRefGoogle Scholar
  21. Irena R, Jacek P, Stanislaw B (2006) Isolation and properties of Aspergillus niger IBT-90 xylanase for bakery. Appl Microbiol Biotechnol 69:665–671CrossRefGoogle Scholar
  22. Kormelink FJM, Voragen AG (1993) Degradation of different [(glucurono) arabino] xylans by a combination of purified xylan-degrading enzymes. Appl Microbiol Biotechnol 38:688–695CrossRefGoogle Scholar
  23. Krisana A, Rutchadaporn S, Jarupan G, Lily E, Sutipa T, Kanyawim K (2005) Endo-1, 4-β-xylanase B from Aspergillus cf. niger BCC14405 isolated in Thailand: purification, characterization and gene isolation. J Biochem Mol Biol 38:17–23Google Scholar
  24. Laemmli UK (1970) Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227:680–685CrossRefGoogle Scholar
  25. Lee CC, Smith M, Kibblewhite-Accinelli RE, Williams TG, Wagschal K, Robertson GH, Wong DW (2006a) Isolation and characterization of a cold-active xylanase enzyme from Flavobacterium sp. Curr Microbiol 52:112–116CrossRefGoogle Scholar
  26. Lee CC, Kibblewhite-Accinelli RE, Wagschal K, Robertson GH, Wong DW (2006b) Cloning and characterization of a cold-active xylanase enzyme from an environmental DNA library. Extremophiles 10:295–300CrossRefGoogle Scholar
  27. Li N, Meng K, Wang Y, Shi P, Luo H, Bai Y, Yang P, Yao B (2008) Cloning, expression, and characterization of a new xylanase with broad temperature adaptability from Streptomyces sp. S9. Appl Microbiol Biotechnol 80:231–240CrossRefGoogle Scholar
  28. Liu R, Qu Y, Jiang Y, Gao P (1999) Purification and characterization of alkaline xylanases from Pseudomonas G6–2. Wei Sheng Wu Xue Bao 39:132–136Google Scholar
  29. Miller GL, Blum R, Glennon WE, Burton AL (1960) Measurement of carboxymethyl cellulase activity. Anal Biochem 2:127–132CrossRefGoogle Scholar
  30. Morosoli R, Bertrand JL, Mondou F, Shareck F, Kluepfel D (1986) Purification and properties of a xylanase from Streptomyces lividans. Biochem J 239:587–592Google Scholar
  31. Petrescu I, Lamotte-Brasseur J, Chessa JP, Ntarima P, Claeyssens M, Devreese B, Marino G, Gerday C (2000) Xylanase from the psychrophilic yeast Cryptococcus adeliae. Extremophiles 4:137–144CrossRefGoogle Scholar
  32. Romanenko LA, Zhukova NV, Rohde M, Lysenko AM, Mikhailov VV, Stackebrandt E (2003) Glaciecola mesophila sp. nov., a novel marine agar-digesting bacterium. Int J Syst Evol Microbiol 53:647–651CrossRefGoogle Scholar
  33. Ruiz-Arribas A, Fernández-Abalos JM, Sánchez P, Garda AL, Santamariá RI (1995) Overproduction, purification, and biochemical characterization of a xylanase (Xys1) from Streptomyces halstedii JM8. Appl Environ Microbiol 61:2414–2419Google Scholar
  34. Saito H, Miura K (1963) Preparation of transforming deoxyribonucleic acid by phenol treatment. Biochim Biophys Acta 72:619–629CrossRefGoogle Scholar
  35. Turkiewiz M, Kalinowska H, Zielinska M, Bielecki S (2000) Purification and characterisation of two endo-1, 4-xylanases from Antarctic krill, Euphasia superba Dana. Comp Biol Physiol Part B 127:325–335CrossRefGoogle Scholar
  36. Wu S, Liu B, Zhang X (2006) Characterization of a recombinant thermostable xylanase from deep-sea thermophilic Geobacillus sp. MT-1 in East Pacific. Appl Microbiol Biotechnol 72:1210–1216CrossRefGoogle Scholar
  37. Yamaura I, Koga T, Matsumoto T, Kato T (1997) Purification and some properties of endo-1, 4-β-D-xylanase from a fresh-water mollusc, Pomacea insularus (de Ordigny). Biosci Biotechnol Biochem 61:615–620CrossRefGoogle Scholar
  38. Yuan KP, Vrijmoed LL, Feng MG (2005) Survey of coastal mangrove fungi for xylanase production and optimized culture and assay conditions. Wei Sheng Wu Xue Bao 45(1):91–96Google Scholar
  39. Zhang G, Huang J, Huang G, Ma L, Zhang X (2007) Molecular cloning and heterologous expression of a new xylanase gene from Plectosphaerella cucumerina. Appl Microbiol Biotechnol 74:339–346CrossRefGoogle Scholar

Copyright information

© Springer-Verlag 2009

Authors and Affiliations

  • Bing Guo
    • 1
  • Xiu-Lan Chen
    • 1
  • Cai-Yun Sun
    • 1
  • Bai-Cheng Zhou
    • 1
  • Yu-Zhong Zhang
    • 1
  1. 1.The State Key Lab of Microbial Technology, Marine Biotechnology Research CenterShandong UniversityJinanChina

Personalised recommendations