Applied Biochemistry and Biotechnology

, Volume 98, Issue 1–9, pp 273–287 | Cite as

Exploration of cellulose surface-binding properties of Acidothermus cellulolyticus Cel5A by site-specific mutagenesis

  • Suzanne L. McCarter
  • William S. Adney
  • Todd B. Vinzant
  • Edward Jennings
  • Fannie Posey Eddy
  • Stephen R. Decker
  • John O. Baker
  • Joshua Sakon
  • Michael E. Himmel
Article

Abstract

Understanding the interactions between cellulases and cellulosic substrates is critical to the development of an efficient artificial cellulase system for conversion of biomass to sugars. We directed specific mutations to the interactive surface of the Acidothermus cellulolyticus EI endoglucanase catalytic domain. The cellulose-binding domain is not translated in these mutants. Amino acid mutations were designed either to change the surface charge of the protein or to modify the potential for hydrogen bonding with cellulose. The relationship between cellulase-to-cellulose (Avicel PH101) binding and hydrolysis activity was determined for various groupings of mutations. While a significant increase in hydrolysis activity was not observed, certain clusters of residues did significantly alter substrate binding and some interesting correlations emerged. In the future, these observations may be used to aid the design of endoglucanases with improved performance on pretreated biomass.

Index Entries

Cellulase endoglucanase site-directed mutagenesis Acidothermus cellulolyticus Cel5A 

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References

  1. 1.
    Wooley, R. and Ruth, M. (1999), Oral Abstract 6-05, Proceedings of the 21st Symposium on Biotechnology for Fuels and Chemicals, National Rewable Energy Labortory, Golden, CO.Google Scholar
  2. 2.
    Sheehan, J. and Himmel, M. E. (1999), Biotechnol. Prog. 15(3), 817–827.CrossRefGoogle Scholar
  3. 3.
    Rubingh, D. N. (1997), Curr. Opin. Biotechnol. 8, 417–422.CrossRefGoogle Scholar
  4. 4.
    Himmel, M. E., Ruth, M. F., and Wyman, C. E. (1999), Curr. Opin. Biotechnol. 10, 358–364.CrossRefGoogle Scholar
  5. 5.
    Henrissat, B. (1994), Cellulose 1, 169–196.CrossRefGoogle Scholar
  6. 6.
    Mohagheghi, A., Grohmann, K., Himmel, M. E., Leighton, L., and Updegraff, D. (1986), Int. J. Syst. Bacteriol. 36, 435–443.Google Scholar
  7. 7.
    Himmel, M. E., Adney, W. S., Grohmann, K., and Tucker, M. P. (1994), US patent no. 5,275,944.Google Scholar
  8. 8.
    Sinnott, M. L. (1990), Chem. Rev. 90, 1171–1202.CrossRefGoogle Scholar
  9. 9.
    Henrissat, B. and Bairoch, A. (1993), Biochem. J. 293, 781–788.Google Scholar
  10. 10.
    Thomas, S. R., Adney, W. S., Baker, J. O., Chou, Y.-C., and Himmel, M. E. (1997), US patent 5,712,142.Google Scholar
  11. 11.
    Sakon, J., Adney, W. S., Himmel, M. E., Thomas, S. R., and Karplus, P. A. (1996), Biochemistry 35, 10,648–10,660.CrossRefGoogle Scholar
  12. 12.
    Wang, Q., Tull, D., Meinke, A., Gilkes, N. R., Warren, R. A. J., Aebersold, R., and Withers, S. G (1993), J. Biol. Chem. 268, 14,096–14,102.Google Scholar
  13. 13.
    Bortoli-German, I., Haiech, J., Chippaux, M., and Barras, F. (1995), J. Mol. Biol. 246, 82–94.CrossRefGoogle Scholar
  14. 14.
    Zhang, S. and Wilson, D. B. (1997), J. Biotech. 57, 101–113.CrossRefGoogle Scholar
  15. 15.
    Zhang, S., Barr, B. K., and Wilson, D. B. (2000), Eur. J. Biochem. 267, 244–252.CrossRefGoogle Scholar
  16. 16.
    Irwin, D. C., Spezio, M., Walker, L. P., and Wilson, D. B. (1993), Biotechnol. Bioeng. 42, 1002–1013.CrossRefGoogle Scholar
  17. 17.
    Tomme, P., Boraston, A., McLean, B., Kormos, J., Creagh, A. L., Sturch, K., Gilkes, N. R., Haynes, C. A., Warren, R. A. J., and Kilburn, D. G. (1998), J. Chromatogr. 715, 283–296.CrossRefGoogle Scholar
  18. 18.
    Reinikainen, T., Ruohonen, L., Nevanen, T., Laaksonen, L., Kraulis, P., Jones, A., Knowles, J. K. C. and Teeri, T. T. (1992), Proteins: Struct., Funct., Genet. 14, 475–482.CrossRefGoogle Scholar
  19. 19.
    Goldstein, M. A. and Doi, R. I. (1994), J. Bacteriol. 176, 7328–7334.Google Scholar
  20. 20.
    Linder, M., Lindeberg, G., Reinikainen, T., Teeri, T. T., and Pettersson, G. (1995), FEBS Lett. 372, 96–98.CrossRefGoogle Scholar
  21. 21.
    Nidetzky, B. and Claeyssens, M. (1994), Biotechnol. Bioeng 44, 961–966.CrossRefGoogle Scholar
  22. 22.
    Kim, D. W., Jang, Y. H., and Jeong, Y. K. (1997), Biotechnol. Lett. 19, 893–897.CrossRefGoogle Scholar
  23. 23.
    Bothwell, M. K., Wilson, D. B., and Walker, L. P. (1997), Enzyme Microb. Technol. 20, 411–417.CrossRefGoogle Scholar
  24. 24.
    Medve, J., Stahlberg, J., and Tjerneld, F. (1997), Appl. Biochem. Biotechnol. 66, 39–56.Google Scholar
  25. 25.
    Bothwell, M. K. and Walker, L. P. (1995), Bioresour. Technol. 53, 21–29.CrossRefGoogle Scholar
  26. 26.
    Lee, I., Evans, B. R., Lane, L. M., and Woodward, J. (1996), Bioresour. Technol. 58, 163–169.CrossRefGoogle Scholar
  27. 27.
    Doner, L. W. and Irwin, P. L. (1992), Analyt. Biochem. 202, 50–53.CrossRefGoogle Scholar
  28. 28.
    Karplus, P. A. (1996), Protein Sci. 5, 1406–142.Google Scholar
  29. 29.
    Hayn, M., Steiner, W., Klinger, R., Steinmuller, H., Sinner, M., and Esterbauer, H. (1993), in Bioconversion of Forest & Agricultural Plant Residues, Saddler, J. N., ed., CAB International, Wallingford, Oxon, UK, pp. 33–72.Google Scholar
  30. 30.
    Tatsumoto, K., Baker, J. O., Tucker, M. P., Oh, K. K., Mohagheghi, A., Grohmann, K., and Himmel, M. E. (1988), Appl. Biochem. Biotechnol. 18, 159–174.Google Scholar
  31. 31.
    Ooshima, H., Burns, D. S., and Converse, A. O. (1990), Biotechnol. Bioeng. 36, 446–452.CrossRefGoogle Scholar
  32. 32.
    Chang, V. S. and Holtzapple, M. T. (2000), Appl. Biochem. Biotechnol. 84–86, 5–37.CrossRefGoogle Scholar
  33. 33.
    Czirnich, W. and Patt, R. (1976), Holzforschung 30, 124–132.Google Scholar
  34. 34.
    Hoffmann, P. and Patt, R. (1976), Holzforschung 30, 19–27.Google Scholar
  35. 35.
    Immergut, E. H. (1975), in The Chemistry of Wood, Browning, B. L., ed., Robert E. Krieger, Huntington, NY, pp. 103–190.Google Scholar

Copyright information

© Humana Press Inc. 2002

Authors and Affiliations

  • Suzanne L. McCarter
    • 1
  • William S. Adney
    • 2
  • Todd B. Vinzant
    • 1
  • Edward Jennings
    • 1
  • Fannie Posey Eddy
    • 1
  • Stephen R. Decker
    • 1
  • John O. Baker
    • 1
  • Joshua Sakon
    • 1
  • Michael E. Himmel
    • 1
  1. 1.Biotechnology for Fuels and Chemicals DivisionNational Bioenergy Center, National Renewable Energy LaboratoryGolden
  2. 2.Department of Chemistry and BiochemistryUniversity of ArkansasFayetteville

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