CLONING AND EXPRESSION OF FULL-LENGTH TRICHODERMA-REESEI CELLOBIOHYDROLASE-I CDNAS IN ESCHERICHIA-COLI

The process of converting lignocellulosic biomass to ethanol via ferme ntation depends on developing economic sources of cellulases. Trichode rma reesei cellobiohydrolase (CBH) I is a key enzyme in the fungal cel lulase system; however, specific process application requirements make modification of the enzyme by site-directed mutagenesis (SDM) an attr active 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 GI724 as a C-terminal fusion to thermostable thioredoxin protein; Full-length exp ression of CBH I was subsequently verified by molecular weight, Wester n blot analysis, and activity on soluble substrates.