Method for the enzymatic synthesis of 2-phospho-d-glycerate from adenosine 5′-triphosphate and d-glycerate via d-glycerate-2-kinase

An enzymatic synthesis of the glycolytic intermediate, 2-phospho-d-glycerate, is reported. This synthesis was undertaken as a means to make 2-phosho-d-glycerate quickly and inexpensively, because this compound is no longer available commercially. The synthesis uses the recombinant enzyme, d-glycerate-2-kinase, and the starting compounds adenosine 5′-triphosphate and d-glycerate from racemic d-l-glycerate. This enzymatic reaction is both stereospecific and regiospecific, producing 2-phospho-d-glycerate exclusively. The product is purified by anion-exchange chromatography using gradient elution with a volatile buffer. This straightforward method produces the potassium salt of 2-phosho-d-glycerate in high yield. The expression and purification of d-glycerate-2-kinase also are discussed.