Journal
BIOTECHNOLOGY PROGRESS
Volume 32, Issue 1, Pages 66-73Publisher
WILEY
DOI: 10.1002/btpr.2197
Keywords
isobutanol; cell free; in vitro; biofuels; alcohol production
Funding
- Department of Energy [DE/SC0006520]
- U.S. Department of Energy (DOE) [DE-SC0006520] Funding Source: U.S. Department of Energy (DOE)
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Producing fuels and chemical intermediates with cell cultures is severely limited by low product concentrations (0.2%(v/v)) due to feedback inhibition, cell instability, and lack of economical product recovery processes. We have developed an alternate simplified production scheme based on a cell-free immobilized enzyme system. Two immobilized enzymes (keto-acid decarboxylase (KdcA) and alcohol dehydrogenase (ADH)) and one enzyme in solution (formate dehydrogenase (FDH) for NADH recycle) produced isobutanol titers 8 to 20 times higher than the highest reported titers with S. cerevisiae on a mol/mol basis. These high conversion rates and low protein leaching were achieved by covalent immobilization of enzymes (ADH) and enzyme fusions (fKdcA) on methacrylate resin. The new enzyme system without in situ removal of isobutanol achieved a 55% conversion of ketoisovaleric acid to isobutanol at a concentration of 0.135 (mole isobutanol produced for each mole ketoisovaleric acid consumed). Further increasing titer will require continuous removal of the isobutanol using an in situ recovery system. (c) 2015 American Institute of Chemical Engineers Biotechnol. Prog., 32:66-73, 2016
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