Monday, May 4, 2009
5-85
Over-expression and purification of Trichoderma reesei glycosyl hydrolases in Pichia and in T. reesei
John S. Scott-Craig, Goutami Banerjee, Melissa S. Borrusch, Subashini Nagendran, and Jonathan D. Walton. MSU-DOE Plant Research Laboratory, Michigan State University, East Lansing, MI 48824
The high cost of degradative enzymes is a limiting factor in the economical conversion of lignocellulose to fermentable sugars. All current biomass-degrading commercial enzyme mixtures are derived from a handful of Ascomyceteous fungi, especially Trichoderma reesei and Aspergillus niger, but the enzymatic activities and relative proportions of current enzyme mixtures are poorly defined. The goal of this project is to produce a defined enzyme mixture that is optimized for pretreated corn stover. On the basis of abundance and predicted importance, twenty enzymes from a proteomics analysis of T. reesei were selected to be expressed either in a heterologous host (Pichia pastoris) or in T. reesei itself. Several expression vectors for T. reesei have been constructed that use either a strong constitutive promoter or a strong cellulose-inducible promoter from T. reesei. The long-term goal is to develop an optimized set of enzymes that has higher specific activity on real lignocellulosic materials than current industrial enzymes. These experiments will also help us identify the key enzymes that should be targets of improvement and further research.