UDP-glucose pyrophosphorylase. Stereochemical course of the reaction of glucose 1-phosphate with uridine-5'[1-thiotriphosphate].

Of the two diastereoisomers of uridine-5'[1-thiotriphosphate] differing in configuration at Palpha, UDP-glucose pyrophosphorylase catalyzes the reaction of the one designated isomer B (Eckstein, F. (1975) Angew. Chem, Int. Ed. Engl. 14, 160-166) with glucose-1-P to produce uridine-5'[1-thiodiphosphate]glucose. Mild acid hydrolysis of the latter, under conditions known to be minimal for the hydrolysis of aldose phosphates, produces diastereoisomer A of uridine-5'[1-thiodiphosphate]. Therefore, the reaction of uridine-5'[1-thiotriphosphate](B) with glucose-1-P catalyzed by UDP-glucose pyrophosphorylase occurs with stereochemical inversion at Palpha to produce uridine-5'[1-thiodiphosphate]glucose(A). The 31P NMR spectra of these nucleotides are presented and used to elucidate the stereochemical course of the reaction. The apparent Km exhibited by this enzyme for uridine-5'[1-thiodiphosphate]glucose isomer A at 2 mM pyrophosphate and pH 7.6 is 75 micron, which is only 2.2 times that for UDP-glucose, 35 micron. The maximal velocity for UDP-glucose is 11 times that for uridine-5'[1-thiodiphosphate]glucose isomer A under these conditions.