The Catalytic Mechanism of a Natural Diels–Alderase Revealed in Molecular Detail

The Diels–Alder reaction, a [4 + 2] cycloaddition of a conjugated diene to a dienophile, is one of the most powerful reactions in synthetic chemistry. Biocatalysts capable of unlocking new and efficient Diels–Alder reactions would have major impact. Here we present a molecular-level description of the reaction mechanism of the spirotetronate cyclase AbyU, an enzyme shown here to be a bona fide natural Diels–Alderase. Using enzyme assays, X-ray crystal structures, and simulations of the reaction in the enzyme, we reveal how linear substrate chains are contorted within the AbyU active site to facilitate a transannular pericyclic reaction. This study provides compelling evidence for the existence of a natural enzyme evolved to catalyze a Diels–Alder reaction and shows how catalysis is achieved.