Direct Epoxidation in Candida antarctica Lipase B Studied by Experiment and Theory

• Candida antarctica lipase B (CALB) is a promiscuous serine hydrolase that, besides its native function, catalyzes different side reactions, such as direct epoxidation. A single-point mutant of CALB demonstrated a direct epoxidation reaction mechanism for the epoxidation of alpha,beta-unsaturated aldehydes by hydrogen peroxide in aqueous and organic solution. Mutation of the catalytically active Ser105 to alanine made the previously assumed indirect epoxidation reaction mechanism impossible. Gibbs free energies, activation parameters, and substrate selectivities were determined both computationally and experimentally. The energetics and mechanism for the direct epoxidation in CALB Ser105Ala were investigated that the reaction proceeds through a two step-mechanism with formation of an oxyanionic intermediate. The active-site residue His224 functions as a general acid-base catalyst with support from Asp187. Oxyanion stabilization is facilitated by two hydrogen bonds from Thr40.

Nyckelord

catalytic promiscuity

enzyme catalysis

epoxidation

hydrolases

molecular dynamics

HYDROGEN-PEROXIDE

ACTIVE-SITE

PEROXYCARBOXYLIC ACIDS

EFFICIENT GENERATION

AM1-BCC MODEL

CATALYSIS

ALKENES

ENZYMES

OXIDATION

CHLOROPEROXIDASE

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