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EN
A change in the dinucleotide reactant from NAD+ to the more oxidizing APAD+ in the malic enzyme reaction results in a change in the mechanism of oxidative decarboxylation of malate from stepwise to concerted [7]. In order to determine whether this is a phenomenon general to metal ion dependent â-hydroxyacid oxidative decarboxylases, tartrate dehydrogenase, which catalyzes a reaction diastereotopic to malic enzyme, was studied using the technique of multiple isotope effects. A primary deuterium isotope effect of 1.41 on V/Kmalate was measured, as well as a primary 13C-isotope effect of 1.0096. A decrease in the measured value of the 13C-isotope effect to 1.0078 is consistent with a stepwise mechanism, as observed for malic enzyme. The 13C-isotope effect with thioNAD+ also decreases from a value of 1.0053 using D-malate to 1.0009 using D-malate-2-D, consistent with stepwise oxidative decarboxylation with this alternative dinucleotide substrate. The data suggest that the change
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