Applied Biochemistry and Microbiology

, Volume 52, Issue 3, pp 277–286 | Cite as

Cloning, purification and characterization of a cellulase-free xylanase from Geobacillus thermodenitrificans AK53

  • M. Irfan
  • H. I. Guler
  • A. O. Belduz
  • A. A. Shah
  • S. Canakci
Article

Abstract

Geobacillus thermodenitrificans AK53 xyl gene encoding xylanase was isolated, cloned and expressed in Escherichia coli. After purifying recombinant xylanase from G. thermodenitrificans AK53 (GthAK53Xyl) to homogeneity by ammonium sulfate precipitation and ion exchange chromatography, biochemical properties of the enzyme were determined. The kinetic studies for GthAK53Xyl showed KM value to be 4.34 mg/mL (for D-xylose) and Vmax value to be 2028.9 μmoles mg–1 min–1. The optimal temperature and pH for enzyme activity were found out to be 70°C and 5.0, respectively. The expressed protein showed the highest sequence similarity with the xylanases of G. thermodenitrificans JK1 (JN209933) and G. thermodenitrificans T-2 (EU599644). Metal cations Mg2+ and Mn2+ were found to be required for the enzyme activity, however, Co2+, Hg2+, Fe2+ and Cu2+ ions caused inhibitor effect on it. GthAK53Xyl had no cellulolytic activity and degraded xylan in an endo-fashion. The action of the enzyme on xylan from oat spelt produced xylobiose and xylopentose. The reported results are suggestive of a xylanase exhibiting desirable kinetics, stability parameters and metal resistance required for the efficient production of xylobiose at industrial scale.

Keywords

xylanase Geobacillus thermodinitrificans thermostable GthAK53Xyl (recombinant xylanase) 

Preview

Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.

References

  1. 1.
    Collins, T., Gerday, C., and Feller, G., FEMS Microbiol. Rev., 2005, vol. 29, no. 1, pp. 3–23.CrossRefPubMedGoogle Scholar
  2. 2.
    Ravalason, H., Gwénaël, J., Daniel, M., Maryvonne, P., and Coutinho, P.M., Appl. Microbiol. Biotechnol., 2008, vol. 80, no. 4, pp. 719–733.CrossRefPubMedGoogle Scholar
  3. 3.
    Wong, K., Tan, L., and Saddler, J.N., Microbiol. Mol. Biol. Rev., 1988, vol. 52, no. 3, pp. 305–317.Google Scholar
  4. 4.
    Beg, Q., Kapoor, M., Mahajan L., and Hoondal, G., Appl. Microbiol. Biotechnol., 2001, vol. 56, no. 3, pp. 326–338.CrossRefPubMedGoogle Scholar
  5. 5.
    Zheng, H.C., Sun, M.Z., Meng, L.C., Pei, H.S., Zhang, X.Q., et al., J. Microbiol. Biotechnol., 2014, vol. 24, no. 4, pp. 489–496.CrossRefPubMedGoogle Scholar
  6. 6.
    Saitou, N. and Nei, M., Mol. Biol. Evol., 1987, vol. 4, no. 4, pp. 406–425.PubMedGoogle Scholar
  7. 7.
    Karaoglu, H., Yanmi, D., Sal, F.A., Celik, A., Canakci, S., and Belduz, A.O., J. Mol. Catal. B: Enzym., 2013, vol. 97, pp. 215–224.CrossRefGoogle Scholar
  8. 8.
    Bergey’s Manual of Systematic Bacteriology, 9th ed., Hensyl, W.M., Ed., Philadelphia, USA: Williams and Wilkins, 1994.Google Scholar
  9. 9.
    Laemmli, U.K., Nature, 1970, vol. 227, no. 5259, pp. 680–685.CrossRefPubMedGoogle Scholar
  10. 10.
    Miller, G.R., Anal. Chem., 1959, vol. 3, pp. 426–428.CrossRefGoogle Scholar
  11. 11.
    Bradford, M.M., Anal. Biochem., 1976, vol. 72, no. 2, pp. 248–254.CrossRefPubMedGoogle Scholar
  12. 12.
    Sharma, A., Adhikari, S., and Satyanarayana, T., World J. Microbiol. Biotechnol., 2007, vol. 23, no. 4, pp. 483–490.CrossRefGoogle Scholar
  13. 13.
    Gerasimova, J., and Kuisiene, N., Microbiology, 2012, vol. 8, no. 4, pp. 418–424.CrossRefGoogle Scholar
  14. 14.
    Verma, D., Anand, A., and Satyanarayana, T., Appl. Biochem. Biotechnol., 2013, vol. 170, no. 1, pp. 119–130.CrossRefPubMedGoogle Scholar
  15. 15.
    Bhosale, S.H., Rao, M.B., and Deshpande, V.V., Microbiol. Rev., 1996, vol. 60, pp. 280–300.PubMedPubMedCentralGoogle Scholar
  16. 16.
    Son-Ng, I., Li, C.W., Yeh, Y., Chen, P.T., Chir, J.L., and Ma, C.H., Extremophiles, 2009, vol. 13, no. 3, pp. 425–435.CrossRefGoogle Scholar
  17. 17.
    Cheng, Y.F., Yang, C.H., and Liu, W.H., Enzyme Microbiol. Technol., 2005, vol. 37, no. 5, pp. 541–546.CrossRefGoogle Scholar
  18. 18.
    Polizeli, M.L., Rizzatti, A.C., Monti, R., Terenzi, H.F, Jorge, J.A., and Amorim, D.S., Appl. Microbiol. Biotechnol., 2005, vol. 67, no. 5, pp. 577–591.CrossRefPubMedGoogle Scholar
  19. 19.
    Knob, A., Beitel, S.M., Fortkamp, D., Terrasan, C.R.F., Fernando de Almeida, A., Biomed. Res. Int., 2013, vol. 8, pp. 728–735.Google Scholar
  20. 20.
    Bastawde, K.B., World. J. Microbiol. Biotechnol., 1992, vol. 8, no. 4, pp. 353–368.CrossRefPubMedGoogle Scholar
  21. 21.
    Zhang, G.M., Huang, J., Huang, G.R, Ma, L.X, and Zhang, X.E., App. Microbiol. Biotechnol., 2007, vol. 74, no. 2, pp. 339–346.CrossRefGoogle Scholar
  22. 22.
    Yin, L., Lin, H., Chiang, Y., and Jiang, S., J. Agricult. Food Chem., 2010, vol. 58, no. 1, pp. 557–562.CrossRefGoogle Scholar
  23. 23.
    Gupta, S., Bhushan, B., and Hoondal, G.S., J. Appl. Microbiol., 2000, vol. 88, no. 2, pp. 325–334.CrossRefPubMedGoogle Scholar
  24. 24.
    Kamble, R.D. and Jadhav, A.R., Asian Pacific J. Trop. Biomed., 2012, vol. 2, pp. 1790–1797.CrossRefGoogle Scholar
  25. 25.
    Salama, M.A., Ismail, K.M.I., Amany, H.A, Abo, E.L., and Genweely, N.S.I., Int. J. Bot., 2008, vol. 4, no. 1, pp. 41–48.CrossRefGoogle Scholar
  26. 26.
    Lu, F., Lu, M.M., Lu, Z., Bie, X., Zhao, H., and Wang, Y., Bioresour. Technol., 2008, vol. 99, no. 13, pp. 5938–5941.CrossRefPubMedGoogle Scholar
  27. 27.
    Cardoso, O.A.V. and Filho, E.X.F., FEMS Microbiol. Lett., 2003, vol. 223, no. 2, pp. 309–314.CrossRefPubMedGoogle Scholar
  28. 28.
    Wu, S., Liu, B., and Zhang, X., Appl. Microbiol. Biotechnol., 2006, vol. 72, no. 6, pp. 1210–1216.CrossRefPubMedGoogle Scholar
  29. 29.
    Saksono, B. and Sukmarini, L., J. Biosci., 2010, vol. 17, no. 4, pp. 189–197.Google Scholar
  30. 30.
    Canakci, S., Inan, K., Kacagan, M., and Belduz, A.O., J. Microbiol. Biotechnol., 2007, vol. 17, no. 8, pp. 1262–1270.PubMedGoogle Scholar
  31. 31.
    Subramaniyan, S. and Prema, P., FEMS Microbiol. Lett., 2000, vol. 183, no. 1, pp. 1–7.CrossRefPubMedGoogle Scholar
  32. 32.
    Basaran, P., Basaran, N., and Hang, Y.D., World J. Microbiol. Biotechnol., 2000, vol. 16, no. 6, pp. 545–550.CrossRefGoogle Scholar
  33. 33.
    Li, X.R., Xu, H., Xie, J., Yi, Q.F., Li, W., Qiao, D.R., Cao, Y., and Cao, Y., J. Microbiol. Biotechnol., 2014, vol. 24, no. 4, pp. 483–488.CrossRefPubMedGoogle Scholar
  34. 34.
    Balaa, B.A., Brijs, K., Gebruers, K., Vandenhaute, J., Wouters, J., and Housen, I., Bioresour. Technol., 2009, vol. 100, no. 24, pp. 6465–6471.CrossRefPubMedGoogle Scholar
  35. 35.
    Zhou, C., Zhang, M., Wang, Y., Guo, W., Liu, Z., Wang, Y., and Wang, W., African J. Microbiol.Res., 2013, vol. 7, no.16, pp. 1535–1542.Google Scholar

Copyright information

© Pleiades Publishing, Inc. 2016

Authors and Affiliations

  • M. Irfan
    • 1
  • H. I. Guler
    • 2
  • A. O. Belduz
    • 3
  • A. A. Shah
    • 1
  • S. Canakci
    • 3
  1. 1.Departmet of Microbiology, Faculty of Biological SciencesQuaid I Azam UniversityIslamabadPakistan
  2. 2.Department Departmet of Molecular Biology and Genetic, Faculty of SciencesKaradeniz Technical UniversityTrabzonTurkey
  3. 3.Department of Biology, Faculty of SciencesKaradeniz Technical UniversityTrabzonTurkey

Personalised recommendations