Crystal Structure of EstZF172 Catalyzing Stereoselectively ()‑CNDE in Pregabalin Biosynthesis.

Pregabalin has garnered extensive clinical application for the management of neuropathic pain and epilepsy owing to its high efficacy and broad drug concentration range. EstZF172 is a key enzyme in the biosynthesis of pregabalin, capable of stereoselectively catalyzing the production of ()-CCMA from the key intermediate -CNDE. The novel crystal structure of EstZF172 indicates that it contains a highly conserved Ser-Lys-Tyr catalytic triad and belongs to the family VIII carboxylesterases. Molecular docking demonstrates that the steric hindrance presented by residues I159 and F239 plays a crucial role in influencing the binding affinity of the chiral substrate ()-CNDE for the catalytic site. The study provides a structural basis and reference for the stereoselective catalysis of EstZF172 and engineering modification of the key enzyme in the synthesis of pregabalin.