Acceleration of the NAD cyanide adduct reaction by lactate dehydrogenase: the equilibrium binding effect as a measure of the activation of bound NAD.
The binary complex of NAD and lactate dehydrogenase reacts reversibly with cyanide to produce a complex (E X NAD-CN) whose noncovalent interactions are similar to those in the E X NADH complex (where E is one-fourth of the tetrameric dehydrogenase). The reaction apparently is a simple bimolecular nucleophilic addition at the 4 position of the bound nicotinamide ring; viz., cyanide does not bind to the enzyme prior to reaction. The value of the dissociation constant for E X NAD-CN is about 1 X 10(-6) M and is independent of pH over the range of 6-8. The equilibrium constant for the reaction of cyanide with E X NAD is about 400-fold larger than that for the nonenzymic process after a statistical correction. This increment in Ke is accounted for by a 220-fold increase in the rate of the forward enzymic reaction (20 M-1 s-1) as compared with an approximately 2-fold decrease for the reverse process (9 X 10(-5) s-1). Thus, the increased value of the rate constant for bond formation in the enzymic reaction is attributed to an equilibrium binding effect that is translated almost entirely into a rate effect on that step (bond formation). Since the nonenzymic reaction is sensitive to solvent composition, this equilibrium binding effect likely is produced by environmental effects at the nicotinamide/dehydronicotinamide part of the coenzyme binding site on the enzyme.
D000818:Animals; D003486:Cyanides / Q000378:metabolism*; D004284:Dogfish; D004789:Enzyme Activation; D007700:Kinetics; D007770:L-Lactate Dehydrogenase / Q000378:metabolism*; D008433:Mathematics; D008954:Models, Biological; D009243:NAD / Q000378:metabolism*; D009536:Niacinamide / Q000031:analogs & derivatives / Q000378:metabolism; D011485:Protein Binding
D003486:Cyanides; D009243:NAD; D009536:Niacinamide; D007770:L-Lactate Dehydrogenase; C008472:N-methylnicotinamide