Journal
JALA
Volume 16, Issue 1, Pages 17-37Publisher
SAGE PUBLICATIONS INC
DOI: 10.1016/j.jala.2010.04.002
Keywords
Candida antarcita lipase B; transesterification; biodisel; Saccharomyces cerevisiae; one-step-changing resin; resin-supported biocatalyst
Funding
- USDA ARS-Mitsubishi Chemical Corporation Nonfunded Cooperative Agreement [3620-41000-121-07]
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A synthetic Candida antarctica lipase B (CALB) gene open reading frame (ORE) for expression in yeast was constructed, and the lycotoxin-I (Lyt-I) C3 variant gene ORE, potentially to improve the availability of the active enzyme at the surface of the yeast cell, was added in frame with the CALB ORE using an automated PCR assembly and DNA purification protocol on an integrated robotic workcell. Saccharomyces cerevisiae strains expressing CALB protein or CALB Lyt-I fusion protein were first grown on 2% (w/v) glucose, producing 9.3 g/L ethanol during fermentation. The carbon source was switched to galactose for GALI-driven expression, and the CALB and CALB Lyt-1 enzymes expressed were tested for fatty acid ethyl ester (biodiesel) production. The synthetic enzymes catalyzed the formation of fatty acid ethyl esters from ethanol and either corn or soybean oil. It was further demonstrated that a one-step-charging resin, specifically selected for binding to lipase, was capable of covalent attachment of the CALB Lyt-1 enzyme, and that the resin-bound enzyme catalyzed the production of biodiesel. High-level expression of lipase in an ethanologenic yeast strain has the potential to increase the profitability of an integrated biorefinery by combining bioethanol production with coproduction of a low-cost biocatalyst that converts corn oil to biodiesel. (JALA 201 1; I 6:17-37)
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