Skip to main content
SpringerLink
Log in

Purification and Characterization of a Low Molecular Weight Endo-xylanase from Mushroom Termitomyces clypeatus

  • Published:
Applied Biochemistry and Biotechnology Aims and scope Submit manuscript

Abstract

A low molecular weight endo-xylanase (EC 3.2.1.8) was purified from an edible mushroom Termitomyces clypeatus grown in submerged medium with oat spelt xylan. Xylanase was purified to apparent homogeneity by ammonium sulfate fractionation and gel filtration chromatography. Its molecular weight was determined by gel filtration chromatography and sodium dodecyl sulfate-polyacrylamide gel electrophoresis to be 12 kDa. The enzyme was found to be most active at 50°C and pH 5.0, being most stable at pH 6.5. The Km for oat spelt xylan was determined to be 10.4 mg/ml. The specificities of the enzyme was observed to be highly specific towards oat spelt xylan and was inhibited by mercuric chloride (HgCl2), N-bromosuccinimide, and trans-1,2-diaminocyclohexane-N′,N′,N′,N′-tetraacetic acid strongly. The inhibitory action of N-bromosuccinimide on enzyme confirmed the presence of one tryptophan residue in its substrate-binding site. Amino acid analysis for xylanase showed the presence of high amount of hydrophobic serine, glycine, threonine, and alanine residues. The N-terminal sequencing study for the previously purified and characterized 56 kDa xylanolytic amyloglucosidase reveal the presence of 33.30% identity with glucoamylase chain A from Aspergillus awamori. The N-terminal sequence analysis of the present 12 kDa enzyme showed highest similarity (72.22% identity) towards xylanase from Neurospora crassa.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3

Similar content being viewed by others

References

  1. Wong, K. Y. Tan, L. L. & Saddler, J. N. (1988). Microbiological Reviews, 52(3), 305–317.

    CAS  Google Scholar 

  2. Aspinall, G. O., & Mckay, J. E. (1958), In: Journal of the Chemical Society Abstract Part. I, 1059–1066.

  3. Dey, P. M. & Brinson, K. (1984). Advances in Carbohydrate Chemistry and Biochemistry, 42, 266–382.

    Article  Google Scholar 

  4. Khowala, S. Mukherjee, M. & Sengupta, S. (1988). Enzyme and Microbial Technology, 10, 563–567.

    Article  CAS  Google Scholar 

  5. Du Toit, P. J. Olivier, S. P. & van Biljon, P. L. (1984). Biotechnology and Bioengineering, 26, 1071–1078.

    Article  Google Scholar 

  6. Gilkes, N. R. Henrissat, B. Kilburn, D. G. Miller, R. C. Jr. & Warren, R. A. (1991). Microbiological Reviews, 55, 303–315.

    CAS  Google Scholar 

  7. Davies, G. & Henrissat, B. (1995). Structure, 3(9), 853–859.

    Article  CAS  Google Scholar 

  8. Beg, Q. K. Kapoor, M. Mahajan, L. & Hoondal, G. S. (2001). Applied Microbiology and Biotechnology, 56, 326–338.

    Article  CAS  Google Scholar 

  9. Gao, F. Jiang, Y. Jhou, G. H. & Han, Z. K. (2008). Animal Feed Science and Technology, 142, 173–184.

    Article  CAS  Google Scholar 

  10. Viikari, L. Kantelinen, A. Sundquist, J. & Linko, M. (1994). FEMS Microbiology Reviews, 13(2–3), 335–350.

    Article  CAS  Google Scholar 

  11. Ghosh, A. K. Jana, M. L. Naskar, A. K. Chatterjee, R. & Sengupta, S. (2002). Biotechnology Letters, 24, 1249–1252.

    Article  CAS  Google Scholar 

  12. Butt, M. S. Nadeem, M. T. Ahmad, Z. & Sultan, M. T. (2008). Food Technology and Biotechnology, 46(1), 22–31.

    CAS  Google Scholar 

  13. Puchart, V. Katapodis, P. Biely, P. et al. (1999). Enzyme and Microbial Technology, 24, 355–361.

    Article  CAS  Google Scholar 

  14. Ghosh, A. K. Naskar, A. K. Jana, M. L. Khowala, S. & Sengupta, S. (1995). Biotechnology Progress, 11, 452–456.

    Article  CAS  Google Scholar 

  15. Ghosh, A. K. Naskar, A. K. & Sengupta, S. (1997). Biochimica et Biophysica Acta, 1339(2), 289–296.

    CAS  Google Scholar 

  16. Ghosh, A. K. & Sengupta, S. (1978). Journal of Food Science and Technology, 15, 237–242.

    CAS  Google Scholar 

  17. Bailey, M. J. Biely, P. & Poutanen, K. (1992). Journal of Biotechnology, 23, 257–270.

    Article  CAS  Google Scholar 

  18. Sengupta, S. Jana, M. L. Sengupta, D. & Naskar, A. K. (2000). Applied Microbiology and Biotechnology, 53, 732–735.

    Article  CAS  Google Scholar 

  19. Somogyi, M. (1952). Journal of Biological Chemistry, 195, 19–23.

    CAS  Google Scholar 

  20. Nelson, N. (1944). Journal of Biological Chemistry, 153, 375–380.

    CAS  Google Scholar 

  21. Bergmeyer, H. U. & Bernt, E. (1974). In H. U. Bergmeyer (Ed.), Methods of enzymatic analysis (3, pp. 1205–1215). London: Academic.

    Google Scholar 

  22. Lowry, O. H. Rosebrough, N. J. Farr, A. L. & Randall, R. J. (1951). Journal of Biological Chemistry, 193, 265–275.

    CAS  Google Scholar 

  23. Gabriel, O. (1971). In W. B. Jakoby (Ed.), Methods in enzymology (enzyme purification and related techniques) (Vol. 22, pp. 565–578). New York: Academic.

    Chapter  Google Scholar 

  24. Laemmli, U. K. (1970). Nature, 227, 680–685.

    Article  CAS  Google Scholar 

  25. Spande, T. F. & Witkop, B. (1967). Methods in Enzymology, 11, 498–506.

    Article  CAS  Google Scholar 

  26. Edelhoch, H. (1967). Biochemistry, 6(7), 1948–1954.

    Article  CAS  Google Scholar 

  27. Raghukumar, C. Muraleedharan, U. Gaud, V. R. & Mishra, R. (2004). Journal of Industrial Microbiology and Biotechnology, 31, 433–441.

    Article  CAS  Google Scholar 

  28. Fang, H. Y. Chang, S. M. Hsieh, M. C. & Fang, T. J. (2007). Journal of Molecular Catalysis. B, Enzymatic, 49, 36–42.

    Article  CAS  Google Scholar 

  29. Chidi, S. B. Godana, B. Ncube, I. van Rensburg, E. J. Cronshaw, A. & Abotsi, E. K. (2008). African Journal of Biotechnology, 7(21), 3939–3948.

    CAS  Google Scholar 

  30. Dutta, T. Sengupta, R. Sahoo, R. Sinha Ray, S. Bhattacharjee, A. & Ghosh, S. (2007). Letters in Applied Microbiology, 44, 206–211.

    Article  CAS  Google Scholar 

  31. Katapodis, P. Vrsanska, M. Kekos, D. et al. (2003). Carbohydrate Research, 338, 1881–1890.

    Article  CAS  Google Scholar 

  32. Quinnghe, C. Xiaoyu, Y. Tiangui, N. Cheng, J. & Qiugang, M. (2004). Process Biochemistry, 39, 1561–1566.

    Article  CAS  Google Scholar 

  33. Ujiie, M. Roy, C. & Yaguchi, M. (1991). Applied and Environmental Microbiology, 57(6), 1860–1862.

    CAS  Google Scholar 

  34. Huang, L. Hesu, T. H. & Wey, T. T. (1991). Biochemical Journal, 278, 329–333.

    CAS  Google Scholar 

  35. e Silva, C. H. C. Puls, J. de Sousa, M. V. & Filho, E. X. F. (1999). Revista de Microbiologia, 30, 114–119.

    Google Scholar 

  36. Ninawe, S. Kapoor, M. & Kuhad, R. C. (2008). Bioresource Technology, 99, 1252–1258.

    Article  CAS  Google Scholar 

  37. Senthilkumar, S. R. Ashokkumar, B. Raj, K. C. & Gunasekaran, P. (2004). Biotechnology Letters, 26, 1283–1287.

    Article  CAS  Google Scholar 

  38. Keskar, S. S. Srinivasan, M. C. & Deshpande, V. V. (1989). Biochemical Journal, 261, 49–55.

    CAS  Google Scholar 

  39. Ximenes, F. A. de Sousa, M. V. Puls, J. de Silva, F. G. Jr. & Filho, E. X. F. (1999). Current Microbiology, 38, 18–21.

    Article  CAS  Google Scholar 

  40. Maleej, I. Belhaj, I. Masmoudi, N. F. & Belghith, H. (2008). Applied Biochemistry and Biotechnology, 158, 200–212. doi:10.1007/s12010-008-8317-x.

    Article  CAS  Google Scholar 

  41. Ghanem, N. B. Yusef, H. H. & Mahrouse, H. K. (2000). Bioresource Technology, 73, 113–121.

    Article  CAS  Google Scholar 

  42. Isil, S. & Nilufer, A. (2005). Brazilian Archives Biology and Technology, 48, 187–193.

    CAS  Google Scholar 

  43. Deshpande, V. Hinge, J. & Rao, M. (1990). Biochimica et Biophysica Acta, 1041(2), 172–177.

    CAS  Google Scholar 

  44. Salles, B. C. Cunha, R. B. Fontes, W. de Sousa, M. V. & Filho, E. X. F. (2000). Journal of Biotechnology, 81, 199–204.

    Article  CAS  Google Scholar 

  45. Breccia, J. D. Sineriz, F. Baigori, M. D. Castro, G. R. & Hatti-Kaul, R. (1998). Enzyme and Microbial Technology, 22, 42–49.

    Article  CAS  Google Scholar 

  46. Huang, X. & Miller, W. (1991). Advances in Applied Mathematics, 12, 337–357.

    Article  Google Scholar 

  47. Galagan, J. E. Calvo, S. E. Borkovich, K. A. et al. (2003). Nature, 422(6934), 859–868.

    Article  CAS  Google Scholar 

Download references

Acknowledgements

The authors wish to thank the Director, IICB for financial support from CSIR Network Project No. NWP 0005. Thanks are also due to University Grants Commission for providing financial support. Grateful thanks are due to Dr. R. Majhi, IICB for the help provided during amino acid analysis and N-terminal sequencing.

Author information

Authors and Affiliations

  1. Drug Development/Diagnostics & Biotechnology Division, Indian Institute of Chemical Biology, 4, Raja S. C. Mullick Road, Kolkata, 700032, India

    Dhananjay Soren, Mohanlal Jana & Anil K. Ghosh

  2. Department of Biotechnology, Heritage Institute of Technology, Chowbaga Road, Anandapur, East Kolkata Township, Kolkata, 700107, India

    Subhabrata Sengupta

Authors
  1. Dhananjay Soren
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Mohanlal Jana
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Subhabrata Sengupta
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Anil K. Ghosh
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Anil K. Ghosh.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Soren, D., Jana, M., Sengupta, S. et al. Purification and Characterization of a Low Molecular Weight Endo-xylanase from Mushroom Termitomyces clypeatus . Appl Biochem Biotechnol 162, 373–389 (2010). https://doi.org/10.1007/s12010-009-8763-0

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s12010-009-8763-0

Keywords

Search

Navigation