Improving the specific activity of TrCel7A by Industrial Protein Engineering

Cellulose hydrolysis involves the synergistic activities of three types of enzymes:  exo-glucanases, endo-glucanases and beta-glucosidases.  Exo-glucanase TrCel7A (CBH1) constitutes about 50% of the total protein in a Trichoderma reesei whole cellulase.  Improvement of TrCel7A will likely result directly in a significant reduction of enzyme dose required to hydrolyze biomass substrates.  In particular, TrCel7A activity has been identified as a limiting activity.  We expressed ~3000 single mutation variants of TrCel7A in Trichoderma reesei and screened them for improved thermostability, and improved specific activity on phosphoric acid swollen cellulose and biomass substrates. Here we present the results of screening of TrCel7A combinatorial libraries, which has led to the identification of variants that display improved cellulolytic activity and improved thermal stability.