초록
<P>Recent demands for the production of biofuels from lignocellulose led to an increased interest in engineered cellulases from <I>Trichoderma reesei</I> or other fungal sources. While the methods to generate such mutant cellulases on DNA level are straightforward, there is often a bottleneck in their production since a correct posttranslational processing of these enzymes is needed to obtain highly active enzymes. Their production and subsequent enzymatic analysis in the homologous host <I>T. reesei</I> is, however, often disturbed by the concomitant production of other endogenous cellulases. As a useful alternative, we tested the production of cellulases in <I>T. reesei</I> in a genetic background where cellulase formation has been impaired by deletion of the major cellulase transcriptional activator gene <I>xyr1</I>. Three cellulase genes (<I>cel7a</I>, <I>cel7b</I>, and <I>cel12a</I>) were expressed under the promoter regions of the two highly expressed genes <I>tef1</I> (encoding translation elongation factor 1-alpha) or <I>cdna1</I> (encoding the hypothetical protein Trire2:110879). When cultivated on <SMALL>D</SMALL>-glucose as carbon source, the Δ<I>xyr1</I> strain secreted all three cellulases into the medium. Related to the introduced gene copy number, the <I>cdna1</I> promoter appeared to be superior to the <I>tef1</I> promoter. No signs of proteolysis were detected, and the individual cellulases could be assayed over a background essentially free of other cellulases. Hence this system can be used as a vehicle for rapid and high-throughput testing of cellulase muteins in a homologous background.</P>