Chembiochem. 2026 Jun 26;27(12):e70438. doi: 10.1002/cbic.70438.

ABSTRACT

4-Oxalocrotonate tautomerase (4-OT) is a promising scaffold for asymmetric biocatalysis, but conventional 4-OT-catalyzed aldol reactions focus solely on aldehyde substrate activation. Herein, we report the directed evolution of a tandem-fused 4-OT enzyme that achieves efficient activation of ketone substrates for enantioselective aldol reactions with substituted benzaldehydes. The engineered biocatalyst yields a range of chiral β-hydroxy ketone products with up to 99% yield and moderate enantioselectivity, while enabling access to both target enantiomers. This work expands the substrate promiscuity and reaction scope of 4-OT, laying a foundation for rational design of next-generation 4-OT variants for advanced organic synthesis and industrial biocatalysis.

PMID:42340114 | DOI:10.1002/cbic.70438