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Repurposing "Ene"-Reductase to Isomerase for Enantiodivergent Synthesis of Allenoates
Heli Cheng1, Kun Zhang1, Pen Chang2
1Key Laboratory of Precise Synthesis of Functional Molecules of Zhejiang Province, Department of Chemistry, School of Science and Research Center for Industries of the Future, Westlake University, Hangzhou, Zhejiang Province 310030, P. R. China.
Abstract:
The synthesis of chiral allenoates through enzymatic asymmetric isomerization of achiral alkynoates via proton transfer is a highly desirable yet unachieved transformation, primarily due to significant product inhibition encountered with natural isomerases. To circumvent this, we repurposed flavin-dependent "ene"-reductases (EREDs), traditionally oxidoreductases, for redox-neutral stereoselective 1,3-proton transfer. Directed evolution of an ERED from Galdieria sulphuraria (GsOYE) generated a panel of new-to-nature isomerases proficient in isomerizing both 3-butynoates and 2-butynoates, affording diverse chiral allenoates with excellent enantiodivergence and negligible product inhibition. The resulting allenoates were leveraged in chirality transfer [2 + 2], [3 + 2], and [4 + 2] cycloadditions to create complex polycycles with up to three stereogenic centers. Mechanistic studies revealed that directed evolution yielded GsOYE mutants capable of operating via either one- or two-base mechanisms, a functional divergence rarely seen with small-molecule catalysts. This study expanded the catalytic repertoire of EREDs, establishing a biocatalytic platform for proton transfer catalysis to set axial chirality.
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