Kinetics of coupled gamma-hydroxybutyrate oxidation and D-glucuronate reduction by an NADP+-dependent oxidoreductase.
A highly purified NADP+-dependent oxidoreductase from hamster liver (1) has been found to catalyze a reaction in which the oxidation of gamma-hydroxybutyrate is coupled to the reduction of D-glucuronate. In the coupled reaction, NADP+ is required in catalytic rather than stoichiometric amounts. When the coupled reaction is carried out under limiting conditions for the oxidation of gamma-hydroxybutyrate (i.e. a very low concentration of NADP+ and an inhibitory concentration of NADPH) the rate of gamma-hydroxybutyrate oxidation is determined by the concentration of D-glucuronate and can be stimulated at least 8-fold by D-glucuronate. The kinetics of the oxidation of gamma-hydroxybutyrate have been studied in both the uncoupled and the coupled reaction. In the coupled reaction the reduction of D-glucuronate drives the oxidation of gamma-hydroxybutyrate; the Km for NADP+ is markedly lower than the Km determined in the uncoupled reaction (1.4 X 10(-6) M as compared to 2 X 10(-5) M), and the inhibition by NADPH can be completely overcome. The kinetics of the uncoupled reaction suggest that, unlike many other dehydrogenases, the oxidation of gamma-hydroxybutyrate catalyzed by this enzyme proceeds by a Rapid Equilibrium Random Bi Bi mechanism.
D000818:Animals; D006224:Cricetinae; D005965:Glucuronates / Q000378:metabolism*; D020723:Glucuronic Acid; D006884:Hydroxybutyrate Dehydrogenase / Q000378:metabolism*; D006885:Hydroxybutyrates / Q000378:metabolism*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D009249:NADP / Q000494:pharmacology; D010084:Oxidation-Reduction; D012978:Sodium Oxybate
D005965:Glucuronates; D006885:Hydroxybutyrates; D009249:NADP; D012978:Sodium Oxybate; D020723:Glucuronic Acid; D006884:Hydroxybutyrate Dehydrogenase; C019416:4-hydroxybutyrate dehydrogenase