Substrate activation by malate induced by oxalate in the Ascaris suum NAD-malic enzyme reaction.
Substrate activation of the rate of the NAD-malic enzyme reaction by malate is obtained in the presence but not in the absence of oxalate. The substrate activation is a result of competition between malate and oxalate for the E.NADH complex, with malate binding to the form of the complex unprotonated at an enzyme group with a pK of 4.9 and oxalate binding preferentially to the protonated form. The off-rate for NADH from the E.NADH complex is completely rate limiting when the group with a pK of 4.9 is protonated but is only one of several rate-limiting steps when it is unprotonated [Kiick, D.M., Harris, B.G., & Cook, P.F. (1986) Biochemistry 25, 227]. The competition by malate with oxalate thus results in an overall increase in the off-rate for NADH as a result of binding to the unprotonated form of E.NADH. Consistent with the proposed mechanism, the deuterium isotope effect on V for the nonsubstrate-activating malate concentration range decreases from 1.6 in the absence of oxalate to 1.3 in the presence of a concentration of oxalate equal to its Kii. The rate equation for the oxalate-induced substrate activation by malate is derived and presented in the Appendix. Data are discussed in terms of the overall mechanism of the NAD-malic enzyme.
Park
S H
SH
Department of Biochemistry, Texas College of Osteopathic Medicine, Fort Worth.
Harris
B G
BG
Cook
P F
PF
D000818:Animals; D001200:Ascaris / Q000201:enzymology*; D004789:Enzyme Activation; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000378:metabolism*; D008293:Malates / Q000494:pharmacology*; D008433:Mathematics; D008928:Mitochondria / Q000201:enzymology; D008962:Models, Theoretical; D010070:Oxalates / Q000494:pharmacology*
D008293:Malates; D010070:Oxalates; D008291:Malate Dehydrogenase