Creighton D J, Rose I A
J Biol Chem. 1976 Jan 10;251(1):61-8.
When cod fish muscle oxalacetate decarboxylase catalyzes the decarboxylation of oxalacetate in the presence of NaBH4, L-lactate results from the reduction of enzyme-bound pyruvate. However, D-lactate results when borohydride reduces the binary enzyme-pyruvate complex formed by adding pyruvate from solution, as reported by others. This observation suggests that there are alternate mechanisms for reduction that are either kinetically or sterically determined for the E-pyruvate forms produced in the two directions. In the process of investigating the mechanism of reduction, the cod fish muscle decarboxylase was discovered to be identical with pyruvate kinase. Decarboxylase activity appears to take place at a site which overlaps the phosphoenolpyruvate binding site on this enzyme, as discussed in the following paper. Crystalline rabbit muscle pyruvate kinase also contains significant decarboxylase activity indicating that the two reactions may be structurally related functions. In the presence of K+, orthophosphate, or ATP the rabbit muscle enzyme catalyzes the detritiation of enzyme-bound pyruvate formed during decarboxylation before release of pyruvate from the enzyme, in analogy with the detritiation of pyruvate formed from P-[3-3/]enolpyruvate in the kinase reaction. This observation is consistent with the formation of an enolpyruvate intermediate common to the kinetic pathways of both reactions. Since the decarboxylase reac.tion is completely stereospecific, within the limits of detection, going with retention of configuration, the protonation of the enolpyruvate intermediate is completely determined by the enzyme as is the case with the enolpyruvate intermediate generated from P-enolpyruvate in the kinase reaction.
当鳕鱼肌肉草酰乙酸脱羧酶在NaBH₄存在下催化草酰乙酸脱羧时，L-乳酸是由酶结合的丙酮酸还原产生的。然而，如其他人所报道的，当硼氢化物还原通过从溶液中加入丙酮酸形成的二元酶-丙酮酸复合物时，会产生D-乳酸。这一观察结果表明，对于在两个方向上产生的E-丙酮酸形式，存在由动力学或空间因素决定的交替还原机制。在研究还原机制的过程中，发现鳕鱼肌肉脱羧酶与丙酮酸激酶相同。脱羧酶活性似乎发生在与该酶上磷酸烯醇丙酮酸结合位点重叠的位点，如下一篇论文所述。结晶兔肌肉丙酮酸激酶也具有显著的脱羧酶活性，表明这两个反应可能是结构相关的功能。在K⁺、正磷酸盐或ATP存在下，兔肌肉酶催化在脱羧过程中形成的酶结合丙酮酸在从酶释放丙酮酸之前的氘代作用，这类似于激酶反应中由P-[3-³H]烯醇丙酮酸形成的丙酮酸的氘代作用。这一观察结果与两个反应的动力学途径中共同的烯醇丙酮酸中间体的形成一致。由于脱羧酶反应在检测限度内是完全立体特异性的，构型保持不变，烯醇丙酮酸中间体的质子化完全由酶决定，这与激酶反应中由P-烯醇丙酮酸产生的烯醇丙酮酸中间体的情况相同。
