Pyruvate aldolases in chiral carbon-carbon bond formation

Abstract

A procedure for the preparation of optically pure a-keto-y-hydroxy carboxylic acids through stereospecific aldol addition catalyzed by pyruvate aldolases from the Entner-Doudoroff and the DeLey-Doudoroff glycolytic pathways is described. This highly versatile fragment serves as a precursor for a variety of commonly encountered functionalities, including ß-hydroxy aldehydes and carboxylic acids, a-amino-y-hydroxy carboxylic acids and a, y-dihydroxy carboxylic acids. The protocol described here uses recombinant His6-tagged KDPG aldolase for the synthesis of (S)-4-hydroxy-2-keto-4-(2'-pyridyl)butyrate. A protocol for evaluating enantiomeric excess through formation of the y-lactone of the dithioacetal followed by chiral-phase gas-liquid chromatography is also described. Enzyme expression and enzymatic synthesis can be accomplished in approximately 1 week. The enzymatic aldol addition proceeds in nearly quantitative yields with enantiomeric excesses greater than 99.7%. © 2007 Nature Publishing Group.