Fast Carbon−Carbon Bond Formation by a Promiscuous Lipase

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Abstract
Lipase B from Candida antarctica was redesigned to catalyze the promiscuous reaction of carbon−carbon bond formation. Mutation of the catalytic serine to alanine afforded a mutant that catalyzed Michael additions of 1,3-dicarbonyls to α,β-unsaturated carbonyl compounds at high specific rates, such as 4000 s-1. The enzyme-catalyzed Michael addition reaction followed saturation kinetics and showed substrate inhibition. The designed enzyme showed high rate enhancements with a catalytic proficiency higher than 108, which is on the same level as that observed for enzymes with native substrates.