Effect of nicotinamide adenine dinucleotide on the oxidation-reduction potentials of lipoamide dehydrogenase from pig heart.
The effect of NAD+ on lipoamide dehydrogenase from pig heart was investigated physicochemically. The observed and theoretical oxidation-reduction mid-point potentials for the oxidized lipoamide dehydrogenase (E)/two-electron-reduced lipoamide dehydrogenase (EH2) couple in the presence on NAD+ were -218 mV and -251 mV, respectively, at pH 6.0. Therefore, unexpectedly the mid-point potential of the enzyme became more positive on NAD+ binding. Decreases in the fluorescence lifetime and intensity and increase in the degree of polarization of enzyme-bound FAD were observed in the presence of NAD+. Fluorescence quenching of bound FAD by NAD+ was released by phenobarbital. The results suggest that NAD+ strengthens the intramolecular dynamic interaction between the isoalloxazine moiety and adenine moiety of bound FAD, and so alters the mid-point potential of the enzyme. These findings indicate that NAD+ acts not only as an acceptor of electrons from EH2, but also as an effector in the flavin-disulfide interaction of EH2.
D000818:Animals; D008058:Dihydrolipoamide Dehydrogenase / Q000378:metabolism*; D007700:Kinetics; D009206:Myocardium / Q000201:enzymology*; D009243:NAD / Q000494:pharmacology*; D010084:Oxidation-Reduction; D011199:Potentiometry; D013552:Swine; D013816:Thermodynamics
D009243:NAD; D008058:Dihydrolipoamide Dehydrogenase