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Applied Biochemistry and Biotechnology

, Volume 57, Issue 1, pp 389–397 | Cite as

Cloning and expression of full-lengthTrichoderma reesi cellobiohydrolase I cDNAs inEscherichia coli

  • Robert A. Laymon
  • William S. Adney
  • Ali Mohagheghi
  • Michael E. Himmel
  • Steven R. Thomas
Session 2 Applied Biological Research

Abstract

The process of converting lignocellulosic biomass to ethanol via fermentation depends on developing economic sources of cellulases.Trichoderma reesei cellobiohydrolase (CBH) I is a key enzyme in the fungal cellulase system; however, specific process application requirements make modification of the enzyme by site-directed mutagenesis (SDM) an attractive goal. To undertake SDM investigations, an efficient, cellulase-free host is required. To test the potential ofEscherichia coli as a host, T.reesei CBH I cDNA was expressed inE. coli strain GI 724 as a C-terminal fusion to thermostable thioredoxin protein. Full-length expression of CBH I was subsequently verified by molecular weight, Western blot analysis, and activity on soluble substrates.

Index entries

Cellobiohydrolase I (CBH I) T. reesei cellulases fusion proteins 

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Copyright information

© Humana Press Inc. 1996

Authors and Affiliations

  • Robert A. Laymon
    • 1
  • William S. Adney
    • 1
  • Ali Mohagheghi
    • 1
  • Michael E. Himmel
    • 1
  • Steven R. Thomas
    • 1
  1. 1.Applied Biological Sciences Branch, Alternative Fuels DivisionNational Renewable Energy LaboratoryGolden

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