Cloning and Expression of Full Length Trichoderma reesei Cellobiohydrolase I cDNAs in Escherichia coli

Robert A. Laymon, William S. Adney, Ali Mohagheghi, Michael E. Himmel, Steven R. Thomas

Research output: Contribution to journalArticlepeer-review

37 Scopus Citations

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 of Escherichia coli as a host, T. reesei CBH I cDNA was expressed in E. 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.

Original languageAmerican English
Pages (from-to)389-397
Number of pages9
JournalApplied Biochemistry and Biotechnology - Part A Enzyme Engineering and Biotechnology
Volume57-58
DOIs
StatePublished - 1996

NREL Publication Number

  • NREL/JA-422-7597

Keywords

  • Cellobiohydrolase I (CBHI)
  • Cellulases
  • Fusion proteins
  • T. reesei

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