Substrate‐Dependent Stereospecificity of Tyl‐KR1: An Isolated Polyketide Synthase Ketoreductase Domain from Streptomyces fradiae

The stereospecificity of an enzymatic reaction depends on the way in which a substrate and its enantiomer bind to the active site. These binding modes cannot be easily predicted. We have studied the stereospecificity and stereoselectivity of the ketoreductase domain Tyl‐KR1 of the tylactone polyketide synthase from Streptomyces fradiae by analysing the stereochemical outcome of the reduction of five different keto ester substrates. The absolute configuration of the Tyl‐KR1 reduction products was determined by using vibrational circular dichroism (VCD) spectroscopy combined with quantum chemical calculations. The conversion of only one of the tested substrates, 2‐methyl‐3‐oxovaleric acid N‐acetylcysteamine thioester, afforded the expected anti‐(2R,3R) configuration of the α‐methyl‐β‐hydroxyl ester product, representing the stereochemistry observed for the physiological polyketide product tylactone. For all other substrates, which were modified with respect to the type of ester and/or the chain length (C₄ instead of C₅), the opposite configuration (anti‐(2S,3S)) was obtained with significant enantio‐ and diastereoselectivity. Inversion of both stereocentres suggests completely different binding modes invoked by only minor modifications of the substrate structure.