RESUMO
An attractive strategy for efficiently forming CS bonds is through the use of diazo compounds SH insertion. However, achieving good enantioselective control in this reaction within a biocatalytic system has proven to be challenging. This study aimed to enhance the activity and enantioselectivity of to enable asymmetric SH insertion. The researchers conducted site-saturation mutagenesis (SSM) on 5 amino acid residues located around the iron carbenoid intermediate within a distance of 5 Å, followed by iterative saturation mutagenesis (ISM) of beneficial mutants. Through this process, the beneficial variant VHbSH(P54R/V98W) was identified through screening with 4-(methylmercapto) phenol as the substrate. This variant exhibited up to 4-fold higher catalytic efficiency and 6-fold higher enantioselectivity compared to the wild-type VHb. Computational studies were also conducted to elucidate the detailed mechanism of this asymmetric SH insertion, explaining how active-site residues accelerate this transformation and provide stereocontrol.
