An epoxide hydrolase from Vigna radiata (VrEH2) affords partial enantioconvergence (84% ee) in the enzymatic hydrolysis of racemic p-nitrostyrene oxide (pNSO), mainly due to insufficient regioselectivity for the (S)-enantiomer (rS = alphaS/betaS = 7.3). To improve the (S)-pNSO regioselectivity, a small but smart library of VrEH2 mutants was constructed by substituting each of four key residues lining the substrate binding site with a simplified amino acid alphabet of Val, Asn, Phe, and Trp. Among the mutants, M263N attacked almost exclusively at Calpha in the (S)-epoxide ring with satisfactory regioselectivity (rS = 99.0), without compromising the original high regioselectivity for the (R)-epoxide (rR = 99.0), resulting in near-perfect enantioconvergence (>99% analytical yield, 98% ee). Structural and conformational analysis showed that the introduced Asn263 formed additional hydrogen bonds with the nitro group in substrate, causing a shift in the substrate binding pose. This shift increased the difference in attacking distances between Calpha and Cbeta, leading to an improved regiopreference toward (S)-pNSO and affording near-perfect enantioconvergence.
Title: Synthesis of novel salidroside esters by lipase-mediated acylation with various functional acyl groups Yu HL, Xu JH, Su JH, Lu WY, Lin GQ Ref: J Biosci Bioeng, 106:65, 2008 : PubMed
Salidroside, a natural glycoside, was enzymatically derived for the first time into novel esters using lipase as biocatalyst. The reaction system of glycoside acylation was optimized, and the effect of solvent nature, concentrations of substrate and biocatalyst, and acyl donors' structure on the acylation was studied. In the optimal system, various structures of acyl donors, either natural or unnatural, including short alkyl acyl groups, long chain acyl groups and acyl donors with aryl group were connected to molecular backbone of the glycoside, forming various structures of novel glycoside esters.
