Effect of NADH-X on cytosolic glycerol-3-phosphate dehydrogenase.
At pH 7.05 NADH-X prepared by incubating NADH with glyceraldehyde-3-phosphate dehydrogenase (E.C. 1.2.1.12) was a potent noncompetitive inhibitor, with respect to coenzyme, of NADPH oxidation by pure rabbit muscle cytosolic glycerol-3-phosphate dehydrogenase (E.C. 1.1.1.8) and also a potent inhibitor of NADPH oxidation catalyzed by this enzyme in a rat pancreatic islet cytosolic fraction. It was a much less potent inhibitor of NADPH oxidation catalyzed by this enzyme in a rat liver cytosolic fraction and of NADH oxidation catalyzed by this enzyme from all three sources. Glycerol-3-phosphate dehydrogenase purified from muscle cytosol contains tightly bound NADH-X, NAD, and ADP-ribose, each in amounts of about 0.1 mol per mole of enzyme polypeptide chain. A deproteinized supernatant of this enzyme contained these three ligands and produced the same type of inhibition of the enzyme described above for prepared NADH-X with a Ki, in the reaction with NADPH at pH 7.05, in the range of 0.2 microM with respect to the total concentration of ligands ([ADP-ribose] + [NAD] + [NADH-X] = 0. 2 microM). However, only the NADH-X component could account for the potent inhibition because NAD, ADP-ribose, and the primary acid product (which can be produced from NADH-X) each had a Ki considerably higher than 0.2 microM. Although at pH 7.05 NADH-X inhibited NADPH oxidation considerably more than NADH oxidation, the reverse was the case at pH 7.38. Since the enzyme purified from muscle contains tightly bound NADH-X, NADH-X might become attached to the enzyme in vivo where it could play a role in regulating the ratio of NADH to NADPH oxidation of the enzyme.
Prabhakar
P
P
Department of Pharmacology, Department of Pediatrics, University of Wisconsin Medical School, 1330 University Avenue, Madison, Wisconsin, 53706, USA.
Laboy
J I
JI
Wang
J
J
Budker
T
T
Din
Z Z
ZZ
Chobanian
M
M
Fahien
L A
LA
D000246:Adenosine Diphosphate Ribose / Q000378:metabolism; D000818:Animals; D002851:Chromatography, High Pressure Liquid; D003600:Cytosol / Q000201:enzymology; D003956:Dialysis; D004789:Enzyme Activation; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism; D050536:Glycerol-3-Phosphate Dehydrogenase (NAD+); D005993:Glycerolphosphate Dehydrogenase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007515:Islets of Langerhans / Q000201:enzymology; D007700:Kinetics; D008024:Ligands; D008099:Liver / Q000201:enzymology; D018482:Muscle, Skeletal / Q000201:enzymology; D009243:NAD / Q000378:metabolism* / Q000494:pharmacology; D009249:NADP / Q000378:metabolism*; D010084:Oxidation-Reduction; D011817:Rabbits; D051381:Rats
D008024:Ligands; D009243:NAD; D000246:Adenosine Diphosphate Ribose; D009249:NADP; D005993:Glycerolphosphate Dehydrogenase; D050536:Glycerol-3-Phosphate Dehydrogenase (NAD+); D005987:Glyceraldehyde-3-Phosphate Dehydrogenases