Engineering of a Reductive Aminase to Enable the Synthesis of a Key Intermediate to a CDK 2/4/6 Inhibitor

Biocatalytic reductive amination reactions with reductive aminases (RedAms) are emerging transformations with a high potential value for pharmaceutical synthesis. Here, we report the identification and engineering of a RedAm to catalyze a reductive amination reaction, making a key intermediate in the synthesis of an investigational cyclin-dependent kinase (CDK) inhibitor, using the relatively bulky benzylamine as a nucleophile. The engineered enzyme contains six mutations with respect to the wild-type and displays high productivity at high substrate concentrations (50-fold improved over the wild-type). After the optimized enzyme variant was identified, crystal structures of both the wild-type and mutant enzymes were solved and used to rationalize how structural changes to the RedAm improved its performance under process conditions. Results suggest that mutations affecting both substrate binding and enzyme thermostability contribute to improved enzyme performance. By enabling the multikilogram-scale synthesis of the chiral intermediate, this work highlights the versatility and industrial utility of RedAm-catalyzed reductive amination.