Lactate dehydrogenase ordered enzyme mechanism

A sequential enzyme-catalyzed reaction mechanism in which two substrates react to form two products and in which there is a preferred order in the binding of substrates and release of products. Several enzymes have been reported to have this type of binding mechanism, including alcohol dehydrogenase , carbamate kinase , lactate dehydrogenase , and ribitol dehydrogenase. ... 

Many enzymes that have NAD+ or NADH as a substrate exhibit the sequential ordered mechanism. Consider lactate dehydrogenase, an important enzyme in glucose metabolism (Section 16.1.9). This enzyme reduces pyruvate to lactate while oxidizing NADH to NAD+. 

The raison d etre for an ordered mechanism may be that binding of the coenzyme creates the binding site for the second substrate by inducing a conformational change of the enzyme 23). There is direct evidence for this from recent X-ray diffraction studies of lactate dehydrogenase 24). The conformational change will, however, be manifest kinetically only if it occurs as an elementary step separate from coenzyme binding, thus. 

Figure 11.8 Mechanism of redox reaction catalyzed by NAD dependent lactate dehydrogenase Lactate dehydrogenase (EC 1.1.1.27) is a tetrameric enzyme which catalyzes the reversible redox reaction between L-lactate and pyruvate via ordered kinetic sequence. The hydride ion is transferred to the proR side of the 4 position of NAD. His 195 acts as an acid-base catalyst removing the proton from lactate during oxidation. The active site loop (residues 98-110) carries Argl09 which helps stabilize the transition state during hydride transfer and contacts required for the substrate specificity.

The distinction between ordered and random binding of substrates during ternary complex formation is more difiicult and rarely absolute. For example the reaction of lactate dehydrogenase from pig heart muscle is classified as an enzyme reaction with an ordered mechanism in the reversible reaction... 

Q. This finding eliminates a truly rapid equilibrium random mechanism, for which k and k must be much smaller than fc 4, k-i, k, and k-2, since the two exchange rates must then be equal. In fact, the differences between the two exchange rates show that the dissociation of A and/or P from the ternary complexes must be slow compared with that of B and/or Q, and also slow relative to the interconversions of the ternary complexes (32). This means that in at least one direction of reaction the dissociation of products in the overall reaction is essentially ordered for all these enzymes, the coenzymes dissociating last, as in the preferred pathway mechanism (Section I,B,4). With malate, lactate, and liver alcohol dehydrogenases, the NAD/NADH exchange rate increased to a... 

---