Directed Evolution of a Hydroxylase into a Decarboxylase for Synthesis of 1-Alkenes from Fatty Acids

Abstract

Inspired by the catalytic mechanism of cytochrome P450 enzymes, we have evolved a fatty acid hydroxylase, P450BSβ, into a decarboxylase after four rounds of saturation mutagenesis that now predominantly converts fatty acids to 1-alkenes. The engineered enzyme directly utilizes inexpensive H2O2 to oxidatively decarboxylate C6–C18 saturated fatty acids as well as plant oil-derived unsaturated fatty acids, displaying high reactivity (e.g., 3,700 h–1 of turnover frequency for myristic acid). With in situ H2O2 supplied by glucose oxidase, this enzyme delivers 1-alkenes in a more productive and scalable fashion (up to 99% yield). Our directed evolution efforts present a strategy for developing new fatty acid decarboxylase for downstream biocatalytic applications.