Heterologous expression of Trichoderma reesei exoglucanase (Cel6A) in Pichia pastoris under the control of GAP promoter

Abstract

Cellulolytic microorganisms produce several cellulase enzymes which have different specificities and modes of action. At least three types of cellulase (endo, exo, and β-glucosidase) are involved in the degradation of cellulose. One of them, exo-β-1,4 glucanase or cellobiohydrolase, can release either glucose or cellobiose from ends of cellulose chains. In this study, we tried to express an exoglucanase (Cel6A) gene heterologically in Pichia pastoris. The Cel6A gene was derived from Trichoderma reesei cellobiohydrolase 2 (CBH2). It was synthetically prepared, and codon optimized for best expression in yeast P. pastoris. The gene was placed under the regulation of the GAP promoter. The recombinant plasmid, named pLIPI-TrCel6A, inserted with T. reesei Cel6A (TrCel6A) gene has been integrated into P. pastoris SMD1168H genome, and the recombinant enzyme has been successfully expressed by P. pastoris with a major product showing a molecular size of around 50 kDa.