Semirational Engineering of a Thermostable Carbonyl Reductase for the Precision Synthesis of (2R,3R)-2-Methyl-2-benzyl-3-hydroxycyclopentanone and Its Analogues

2,2-Disubstituted-3-hydroxycyclopentanones are important chiral intermediates for natural products and pharmaceuticals. Through semirational engineering of a thermostable carbonyl reductase CBCR from Cupriavidus sp. BIS7, a mutant L91C/F93I was obtained. Mutant L91C/F93I showed 4- to 36-fold enhanced activities toward 2-methyl-2-benzyl-1,3-cyclopentanedione and its analogues, affording the (2R,3R)-stereoisomers with >99% ee and >99% de. Enzyme-substrate docking studies were performed to reveal the molecular basis for the activity and stereoselectivity improvements.