(528d) Performance and Applications of a Yeast Surface Display-Based Biocatalyst

A whole-cell biocatalyst has been developed using surface display (SD) on recombinant Saccharomyces cerevisiae. Enzymes can be produced by the yeasts and attached to the outer cell surface covalently or non-covalently. Two enzymes have been displayed: lipase B from Candida antartica (CALB) and lipase from Photobacterium lipolyticum M37 (M37L).  To characterize the activity and protein concentration on the cell surface, a flow cytometry method has been developed.  Active enzymes are labeled with a reactive probe that is linked to a detectable dye via a “click” reaction. All proteins produced as part of the SD construct are then labeled via a C-terminal epitope tag.  The amount of active and total protein can be measured in respective flow cytometry fluorescence channels.

The performance of the whole-cell biocatalyst was evaluated in several ways. The cells were lyophilized and assays for hydrolytic and synthetic activity were performed. The assays are based on the release of para-nitrophenol and the formation of propyl laurate, respectively. The catalysts were also used in the synthesis of several commercial products, including glycerol carbonate (a value-added derivative of glycerol).