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Research article

References

  1. Alfonso, C., O. M. Nuero, F. Santamaría, and F. Reyes. 1995. Purification of a heat stable chitin deacetylase from Aspergillus nidulans and its role in cell wall degradation. Curr. Microbiol. 30: 49-54.
    CrossRef
  2. Araki, Y. and E. A. Ito. 1975. Pathway of chitosan formation in Mucor rouxii. Eur. J. Biochem. 55: 71-78.
    CrossRef
  3. Blair, D. E., O. Hekmat, A. W. Schuttelkopf, B. Shrestha, K. Tokuyasu, S. G. Withers, and D. M. F. van Aalten. 2006. Structure and mechanism of chitin deacetylase from the fungal pathogen Colletotrichum lindemuthianum. Biochemistry 45:9416-9426.
    CrossRef
  4. Carr, P. A., J. S. Park, Y. J. Lee, T. Yu, S. Zhang, and J. M. Jacobson. 2004. Protein-mediated error correction for de novo DNA synthesis. Nucleic Acids Res. 32: e162.
    CrossRef
  5. Caufrier, F., A. Martinou, C. Dupont, and V. Bouriotis. 2003. Carbohydrate esterase family 4 enzymes: Substrate specificity. Carbohydr. Res. 338: 687-692.
    CrossRef
  6. Chen, G. Q., I. Choi, B. Ramachandran, and J. E. Gouaux. 1994. Total gene synthesis: Novel single step and convergent strategies applied to the construction of a 779 base pair bacteriorhodopsin gene. J. Am. Chem. Soc. 11: 8799-8800.
    CrossRef
  7. Christodoulidou, A., P. Briza, A. Ellinger, and V. Bouriotis. 1999. Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett. 460: 275-279.
    CrossRef
  8. Dutta, P. K., J. Dutta, and V. S. Tripathi. 2004. Chitin and chitosan: Chemistry, properties and applications. J. Sci. Ind. Res. 63: 20-31.
  9. Gao, X. D., T. Katsumoto, and K. Onodera. 1995. Purification and characterization of chitin deacetylase from Absidia coerulea. J. Biochem. 117: 257-263.
    CrossRef
  10. Gauthier, C., F. Clerisse, J. Dommes, and M. F. Jaspar-Versali. 2008. Characterization and cloning of chitin deacetylases from Rhizopus circinans. Protein Expr. Purif. 59: 127-137.
    CrossRef
  11. Hekmat, O., K. Tokuyasu, and S. G. Withers. 2003. Subsite structure of the endo-type chitin deacetylase from a Deuteromycete, Colletotrichum lindemuthianum: An investigation using steadystate kinetic analysis and MS. Biochem. J. 374: 369-380.
    CrossRef
  12. Jaworska, M. M., J. Bryjak, and J. Liesiene. 2009. A search of an optimal carrier for immobilization of chitin deacetylase. Cellulose 16: 261-270.
    CrossRef
  13. Kafetzopoulos, D., A. Martinou, and V. Bouriotos. 1993. Bioconversion of chitin to chitosan: Purification and characterization of chitin deacetylase from Mucor rouxii. Proc. Natl. Acad. Sci. USA 90: 2564-2568.
    CrossRef
  14. Kauss, H., W. Jeblick, and D. H. Young. 1983. Chitin deacetylase from the plant pathogen Colletotrichum lindemuthianum. Plant Sci. 28: 231-236.
  15. Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680685.
    CrossRef
  16. Matsuo, Y., K. Tanaka, H. Matsuda, and M. Kawamukai. 2005. Cda1+, encoding chitin deacetylase is required for proper spore formation in Schizosaccharomyces pombe. FEBS Lett. 579:2737-2743.
    CrossRef
  17. Shevchenko, A., M. Wilm, O. Vorm, and M. Mann. 1996. Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal. Chem. 68: 850-858.
    CrossRef
  18. Shrestha, B., K. Blondeau, W. F. Stevens, and F. L. Hegarat. 2004. Expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris: Purification and characterization. Protein Expr. Purif. 38: 196-204.
    CrossRef
  19. Tsigos, I. and V. Bouriotis. 1995. Purification and characterization of chitin deacetylase from Colletotrichum lindemuthianum. J. Biol. Chem. 270: 26286-26291.
    CrossRef
  20. Sreekrishna, K., R. G. Brankamp, K. E. Kropp, D. T. Blankenship, J. T. Tsay, P. L. Smith, et al. 1997. Strategies for optimal synthesis and secretion of heterologous proteins in the methylotrophic yeast Pichia pastoris. Gene 190: 55-62.
    CrossRef
  21. Tokuyasu, K., S. Kaneko, K. Hayashi, and Y. Mori. 1999. Production of recombinant chitin deacetylase in the culture medium of Escherichia coli cells. FEBS Lett. 458: 23-26.
    CrossRef
  22. Tokuyasu, K., M. Ohnishi-Kameyama, and K. Hayashi. 1996. Purification and characterization of extracellular chitin deacetylase from Colletotrichum lindemuthianum. Biosci. Biotechnol. Biochem. 60: 1598-1603.
    CrossRef
  23. Tokuyasu, K., M. Ohnishi-Kameyama, K. Hayashi, and Y. Mori. 1999. Cloning and expression of chitin deacetylase gene from a Deuteromycete, Colletotrichum lindemuthianum. J. Biosci. Bioeng. 87: 418-423.
    CrossRef
  24. Tokuyasu, K., M. Mitsutomi, I. Yamaguchi, K. Hayashi, and Y. Mori. 2000. Recognition of chitooligosaccharides and their Nacetyl groups by putative subsites of chitin deacetylase from a Deuteromycete, Colletotrichum lindemuthianum. Biochemistry 39: 8837-8843.
    CrossRef
  25. Tsigos, I., A. Martinou, D. Kafetzopoulos, and V. Bouriotis. 2000. Chitin deacetylases: New, versatile tools in biotechnology. Trends Biotechnol. 18: 305-312.
    CrossRef
  26. Wang, Y., J. Z. Song, Q. Yang, Z. H. Liu, X. M. Huang, and Y. Chen. 2010. Cloning of a heat-stable chitin deacetylase gene from Aspergillus nidulans and its functional expression in Escherichia coli. Appl. Biochem. Biotechnol. 162: 843-854.
    CrossRef
  27. Win, N. N. and W. F. Stevens. 2001. Shrimp chitin as substrate for fungal chitin deacetylase. Appl. Microbiol. Biotechnol. 57:334-341.
    CrossRef
  28. Zhang, G. M., Y. Hu, and Y. H. Zhuang. 2006. Molecular cloning and heterologous expression of an alkaline xylanase from Bacillus pumilus HBP8 in Pichia pastoris. Biocatal. Biotransform. 24: 371-379.
    CrossRef
  29. Zhao, Y., R. D. Park, and R. A. Muzzarelli. 2010. Chitin deacetylases: Properties and applications. Mar. Drugs 8: 24-46.
    CrossRef

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Article

Research article

J. Microbiol. Biotechnol. 2012; 22(9): 1202-1207

Published online September 28, 2012 https://doi.org/10.4014/jmb.1112.12026

Copyright © The Korean Society for Microbiology and Biotechnology.

Synthesis and High Expression of Chitin Deacetylase from Colletotrichum lindemuthianum in Pichia pastoris GS115

Lixin Kang 1, Xiaomei Chen 2, Chao Zhai 1 and Lixin Ma 1*

1Hubei Key Laboratory of Industrial Biotechnology, College of Life Sciences, Hubei University, Wuhan, Hubei Province, 430062, P. R. China, 2State Key Laboratory of Genetic Engineering, School of Life Sciences, Fudan University, Shanghai, 200433, P. R. China

Received: December 12, 2011; Accepted: May 15, 2012

Abstract

A gene, ClCDA, encoding chitin deacetylase from
Colletotrichum lindemuthianum, was optimized according
to the codon usage bias of Pichia pastoris and synthesized
in vitro by overlap extension PCR. It was secretorily
expressed in P. pastoris GS115 using the constitutive expression
vector pHMB905A. The expression level reached the
highest with 110 mg/l culture supernatant after 72 h of
methanol induction, which comprised 77.27 U/mg chitin
deacetylase activity. SDS-PAGE, mass spectrometry, and
deglycosylation assays demonstrated that partial recombinant
protein was glycosylated with an apparent molecular mass
of 33 kDa. The amino acid sequences of recombinant
proteins were confirmed by mass spectrometry.

Keywords: Chitin deacetylase, Colletotrichum lindemuthianum, high-expression, Pichia pastoris, Synthesis

References

  1. Alfonso, C., O. M. Nuero, F. Santamaría, and F. Reyes. 1995. Purification of a heat stable chitin deacetylase from Aspergillus nidulans and its role in cell wall degradation. Curr. Microbiol. 30: 49-54.
    CrossRef
  2. Araki, Y. and E. A. Ito. 1975. Pathway of chitosan formation in Mucor rouxii. Eur. J. Biochem. 55: 71-78.
    CrossRef
  3. Blair, D. E., O. Hekmat, A. W. Schuttelkopf, B. Shrestha, K. Tokuyasu, S. G. Withers, and D. M. F. van Aalten. 2006. Structure and mechanism of chitin deacetylase from the fungal pathogen Colletotrichum lindemuthianum. Biochemistry 45:9416-9426.
    CrossRef
  4. Carr, P. A., J. S. Park, Y. J. Lee, T. Yu, S. Zhang, and J. M. Jacobson. 2004. Protein-mediated error correction for de novo DNA synthesis. Nucleic Acids Res. 32: e162.
    CrossRef
  5. Caufrier, F., A. Martinou, C. Dupont, and V. Bouriotis. 2003. Carbohydrate esterase family 4 enzymes: Substrate specificity. Carbohydr. Res. 338: 687-692.
    CrossRef
  6. Chen, G. Q., I. Choi, B. Ramachandran, and J. E. Gouaux. 1994. Total gene synthesis: Novel single step and convergent strategies applied to the construction of a 779 base pair bacteriorhodopsin gene. J. Am. Chem. Soc. 11: 8799-8800.
    CrossRef
  7. Christodoulidou, A., P. Briza, A. Ellinger, and V. Bouriotis. 1999. Yeast ascospore wall assembly requires two chitin deacetylase isozymes. FEBS Lett. 460: 275-279.
    CrossRef
  8. Dutta, P. K., J. Dutta, and V. S. Tripathi. 2004. Chitin and chitosan: Chemistry, properties and applications. J. Sci. Ind. Res. 63: 20-31.
  9. Gao, X. D., T. Katsumoto, and K. Onodera. 1995. Purification and characterization of chitin deacetylase from Absidia coerulea. J. Biochem. 117: 257-263.
    CrossRef
  10. Gauthier, C., F. Clerisse, J. Dommes, and M. F. Jaspar-Versali. 2008. Characterization and cloning of chitin deacetylases from Rhizopus circinans. Protein Expr. Purif. 59: 127-137.
    CrossRef
  11. Hekmat, O., K. Tokuyasu, and S. G. Withers. 2003. Subsite structure of the endo-type chitin deacetylase from a Deuteromycete, Colletotrichum lindemuthianum: An investigation using steadystate kinetic analysis and MS. Biochem. J. 374: 369-380.
    CrossRef
  12. Jaworska, M. M., J. Bryjak, and J. Liesiene. 2009. A search of an optimal carrier for immobilization of chitin deacetylase. Cellulose 16: 261-270.
    CrossRef
  13. Kafetzopoulos, D., A. Martinou, and V. Bouriotos. 1993. Bioconversion of chitin to chitosan: Purification and characterization of chitin deacetylase from Mucor rouxii. Proc. Natl. Acad. Sci. USA 90: 2564-2568.
    CrossRef
  14. Kauss, H., W. Jeblick, and D. H. Young. 1983. Chitin deacetylase from the plant pathogen Colletotrichum lindemuthianum. Plant Sci. 28: 231-236.
  15. Laemmli, U. K. 1970. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature 227: 680685.
    CrossRef
  16. Matsuo, Y., K. Tanaka, H. Matsuda, and M. Kawamukai. 2005. Cda1+, encoding chitin deacetylase is required for proper spore formation in Schizosaccharomyces pombe. FEBS Lett. 579:2737-2743.
    CrossRef
  17. Shevchenko, A., M. Wilm, O. Vorm, and M. Mann. 1996. Mass spectrometric sequencing of proteins from silver-stained polyacrylamide gels. Anal. Chem. 68: 850-858.
    CrossRef
  18. Shrestha, B., K. Blondeau, W. F. Stevens, and F. L. Hegarat. 2004. Expression of chitin deacetylase from Colletotrichum lindemuthianum in Pichia pastoris: Purification and characterization. Protein Expr. Purif. 38: 196-204.
    CrossRef
  19. Tsigos, I. and V. Bouriotis. 1995. Purification and characterization of chitin deacetylase from Colletotrichum lindemuthianum. J. Biol. Chem. 270: 26286-26291.
    CrossRef
  20. Sreekrishna, K., R. G. Brankamp, K. E. Kropp, D. T. Blankenship, J. T. Tsay, P. L. Smith, et al. 1997. Strategies for optimal synthesis and secretion of heterologous proteins in the methylotrophic yeast Pichia pastoris. Gene 190: 55-62.
    CrossRef
  21. Tokuyasu, K., S. Kaneko, K. Hayashi, and Y. Mori. 1999. Production of recombinant chitin deacetylase in the culture medium of Escherichia coli cells. FEBS Lett. 458: 23-26.
    CrossRef
  22. Tokuyasu, K., M. Ohnishi-Kameyama, and K. Hayashi. 1996. Purification and characterization of extracellular chitin deacetylase from Colletotrichum lindemuthianum. Biosci. Biotechnol. Biochem. 60: 1598-1603.
    CrossRef
  23. Tokuyasu, K., M. Ohnishi-Kameyama, K. Hayashi, and Y. Mori. 1999. Cloning and expression of chitin deacetylase gene from a Deuteromycete, Colletotrichum lindemuthianum. J. Biosci. Bioeng. 87: 418-423.
    CrossRef
  24. Tokuyasu, K., M. Mitsutomi, I. Yamaguchi, K. Hayashi, and Y. Mori. 2000. Recognition of chitooligosaccharides and their Nacetyl groups by putative subsites of chitin deacetylase from a Deuteromycete, Colletotrichum lindemuthianum. Biochemistry 39: 8837-8843.
    CrossRef
  25. Tsigos, I., A. Martinou, D. Kafetzopoulos, and V. Bouriotis. 2000. Chitin deacetylases: New, versatile tools in biotechnology. Trends Biotechnol. 18: 305-312.
    CrossRef
  26. Wang, Y., J. Z. Song, Q. Yang, Z. H. Liu, X. M. Huang, and Y. Chen. 2010. Cloning of a heat-stable chitin deacetylase gene from Aspergillus nidulans and its functional expression in Escherichia coli. Appl. Biochem. Biotechnol. 162: 843-854.
    CrossRef
  27. Win, N. N. and W. F. Stevens. 2001. Shrimp chitin as substrate for fungal chitin deacetylase. Appl. Microbiol. Biotechnol. 57:334-341.
    CrossRef
  28. Zhang, G. M., Y. Hu, and Y. H. Zhuang. 2006. Molecular cloning and heterologous expression of an alkaline xylanase from Bacillus pumilus HBP8 in Pichia pastoris. Biocatal. Biotransform. 24: 371-379.
    CrossRef
  29. Zhao, Y., R. D. Park, and R. A. Muzzarelli. 2010. Chitin deacetylases: Properties and applications. Mar. Drugs 8: 24-46.
    CrossRef