Stereoselective decarboxylation of β-functionalized carboxylic acids enabled by engineered fatty acid photodecarboxylase

Abstract

Here, the light-driven fatty acid photodecarboxylase (CvFAP) was engineered to broaden its substrate spectrum for stereoselective decarboxylation of β-functionalized carboxylic acids, producing chiral carboxylic acids and chiral amines/alcohols. Initially, saturation mutagenesis was performed at V463, a substrate-5-Å-proximal residue identified by AFISHED scanning (AFISHED: seven representative amino acids). Two beneficial variants V463T and V463A were obtained, showing 210% and 150% increases in yield over wild-type (WT) CvFAP in stereoselective decarboxylation of 10 mM mediumchain substrate β-hydroxydecanoic acid. Molecular dynamics simulation and interaction analysis reveal that the distance between the substrate and FAD in above variants is 1.41-1.56 Å shortened than that of WT, due to microperturbationinduced "Hydrogen Bond Network Remodeling". Moreover, clustering analysis reveals that for short-chain substrate βhydroxyhexanoic acid, the (R)-isomer preferentially forms an intramolecular hydrogen bond, facilitating the proper anchoring of the carboxyl group within active site. This mechanism explains the strict enantioselectivity of CvFAP, as evidenced by (S)-2-pentanol in 99% ee achieved by V463A. Furthermore, photostability of CvFAP was proven to be a vital factor influencing its activity. This work expands the catalytic spectrum of CvFAP toward decarboxylation of β-hydroxy carboxylic acids with varying chain lengths, provides a promising strategy for further resolution of β-functionalized carboxylic acids and synthesis of chiral chemicals by utilizing renewable solar energy.