Reversal and Amplification of the Enantioselectivity of Biocatalytic Desymmetrization toward Meso Heterocyclic Dicarboxamides Enabled by Rational Engineering of Amidase

By rational engineering of amidase, the efficient biocatalytic desymmetrization of meso O -heterocyclic dicarboxamides for synthesizing both antipodes of functionalized cyclic motifs was presented. Based on the enzyme-substrate binding model suggested by molecular docking, a rational mutagenesis strategy was established. The reversal and amplification of enantioselectivity of amidase were achieved by generating and testing only 10 variants. This enabled the quick access of both antipodes of products in very good yields and up to 99.5% ee under mild conditions. The engineered biocatalyst exhibits a wide substrate promiscuity and can expand to both N -heterocyclic and carbocyclic dicarboxamides with the retained high efficiency and excellent enantioselectivity. The desymmetrization mechanism for amidase, including the wild-type and the variant, was revealed by molecular dynamics simulations and quantum mechanical/molecular mechanical modeling. It suggests a delicate cooperation between the activation and binding sites by nesting the cyclic skeleton of the substrates.