Computational Design of an Enzyme Catalyst for a Stereoselective Bimolecular Diels-Alder Reaction

TLDR

The Diels‑Alder reaction, a cornerstone of organic synthesis that forms two C–C bonds and up to four stereogenic centers in one step, has no known natural enzymatic catalysts for bimolecular variants. The study aims to de novo computationally design and experimentally characterize enzymes that catalyze a bimolecular Diels‑Alder reaction with high stereoselectivity and substrate specificity. Computational design generated enzyme scaffolds, which were expressed and assayed experimentally to evaluate catalytic activity and stereoselectivity. X‑ray crystallography confirmed that the most active enzyme’s structure matches the design, and binding‑site substitutions reprogrammed substrate specificity, demonstrating that such stereoselective catalysts can be broadly useful in synthetic chemistry.

Abstract

The Diels-Alder reaction is a cornerstone in organic synthesis, forming two carbon-carbon bonds and up to four new stereogenic centers in one step. No naturally occurring enzymes have been shown to catalyze bimolecular Diels-Alder reactions. We describe the de novo computational design and experimental characterization of enzymes catalyzing a bimolecular Diels-Alder reaction with high stereoselectivity and substrate specificity. X-ray crystallography confirms that the structure matches the design for the most active of the enzymes, and binding site substitutions reprogram the substrate specificity. Designed stereoselective catalysts for carbon-carbon bond-forming reactions should be broadly useful in synthetic chemistry.