Enzymatic synthesis of cytidine diphosphate 3,6-dideoxyhexoses. 3. Cytidine diphosphate D-glucose oxidoreductase.

A procedure is described for the purification of CDP-Dglucose oxidoreductase which catalyzes formation of CDP-4keto-6-deoxy-D-glucose and for enzymes El and ES which catalyze the further reduction to CDP-4-keto-3,6-dideoxy-Dglucose. All three enzymes have been purified to homogeneity as judged by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate. CDP-D-glucose oxidoreductase has been shown to consist of two identical subunits of molecular weight of about 43,000 per subunit. Enzymes El and ES consist of simple polypeptide chains of molecular weight of about 61,000 and 41,000, respectively. Preliminary studies on the mechanism of the reaction suggest that enzyme El binds the substrate and that pyridoxamine 5’-phosphate is essential for the binding. Enzyme E, has been shown to possess NADH oxidase activity and may be the reductase. A mechanism involving participation of the essential cofactor, pyridoxamine-5’-phosphate, is discussed