Engineering an Alcohol Dehydrogenase from Kluyveromyces polyspora for Efficient Synthesis of Ibrutinib Intermediate

Abstract ( S )‐ N ‐Boc‐3‐hydroxypiperidine [( S )‐NBHP] is a key intermediate for the synthesis of mantle cell lymphoma drug, ibrutinib. Here, Kp ADH, an alcohol dehydrogenase from Kluyveromyces polyspora , exhibits evolutionary potential in the asymmetric reduction of N ‐Boc‐3‐piperidone (NBPO) to ( S )‐NBHP. By screening key residues in substrate binding pocket of Kp ADH, an excellent variant Y127W was obtained with 6‐fold improved activity of 119.3 U mg −1 , 1.8‐fold enhanced half‐life of 147 h and strict S ‐stereoselectivity (>99% ee ). When catalyzed by Y127W, a complete conversion of 600 g L −1 NBPO was achieved at a substrate to catalyst ratio (S/C) of 30 in 10 h. Based on crystal‐structure of Y127W, molecular docking and dynamic simulations reveal hydrogen bonds formed between W127 and Boc group of NBPO, as well as improved structural stability mainly contribute to the increased catalytic activity and stereoselectivity of Y127W. This study offers guidance for engineering ADHs for biosynthesis of chiral heterocyclic alcohols, and provides insights into mechanisms in catalytic activity and stereoselectivity toward carbonyl‐containing heterocyclic substrates. magnified image