Huhta D W, Heckenthaler T, Alvarez F J, Ermer J, Hübner G, Schellenberger A, Schowen R L
Department of Chemistry, University of Kansas, Lawrence 66045-0046.
Acta Chem Scand (Cph). 1992 Aug;46(8):778-88. doi: 10.3891/acta.chem.scand.46-0778.
Pyruvate decarboxylase (PDC) catalyzes the decarboxylation of pyruvate anion by a factor of around 10(12), compared with the non-enzymic decarboxylation by thiamine, under standard state conditions of 1 mM pyruvate and thiamine diphosphate (TDP), pH 6.2. Free-energy diagrams constructed on the basis of earlier measurements for the enzymic and non-enzymic reactions give some information on catalysis by PDC. PDC stabilizes the reactant state preceding TDP addition to pyruvate by 76 kJ mol-1 and the transition state for the addition by 83 kJ mol-1. PDC stabilizes the reactant state preceding decarboxylation (presumably alpha-lactyl-TDP) by 27 kJ mol-1 and the transition state for decarboxylation by 68 kJ mol-1. In addition, the free-energy diagrams reveal a leveling of reactant-state free energies in the enzymic reaction compared with the non-enzymic reaction, in that the former are nearly equal to each other. The enzyme-bound transition-state energies are similarly leveled. The energetic leveling of reactant states has been noted by Albery, Knowles and their coworkers in many enzymic reactions and termed 'matched internal thermodynamics.' They showed that the result would arise naturally (and inevitably) in the 'evolution to perfection' of enzymes, when the evolutionary process was treated by a deterministic model. The critical assumption of this model was the validity of a Marcus-type or Brønsted-type linear free-energy relationship between rate and equilibrium constants for reactions occurring wholly within enzyme complexes. Here a completely stochastic simulation of molecular evolution, with no deterministic assumptions, is shown to reproduce both 'matched internal thermodynamics' and the 'matched internal kinetics' or leveling of transition-state energies noted here. The Albery-Knowles result is thus more general than might have been supposed.
在1 mM丙酮酸和硫胺二磷酸（TDP）、pH 6.2的标准状态条件下，与硫胺的非酶促脱羧反应相比，丙酮酸脱羧酶（PDC）催化丙酮酸阴离子脱羧的速率提高约10¹²倍。根据早期对酶促反应和非酶促反应的测量构建的自由能图，提供了一些关于PDC催化作用的信息。PDC使丙酮酸添加TDP之前的反应物状态稳定76 kJ/mol，使添加TDP的过渡态稳定83 kJ/mol。PDC使脱羧反应之前的反应物状态（可能是α-乳酰-TDP）稳定27 kJ/mol，使脱羧反应的过渡态稳定68 kJ/mol。此外，自由能图显示，与非酶促反应相比，酶促反应中反应物状态的自由能趋于平缓，因为前者彼此几乎相等。酶结合的过渡态能量也同样趋于平缓。Albery、Knowles及其同事在许多酶促反应中都注意到了反应物状态的能量平缓现象，并将其称为“匹配的内部热力学”。他们表明，当用确定性模型处理进化过程时，在酶的“进化至完美”过程中，这一结果会自然（且不可避免地）出现。该模型的关键假设是，对于完全在酶复合物内发生的反应，速率和平衡常数之间存在Marcus型或Brønsted型线性自由能关系。在此，展示了一个完全随机的分子进化模拟，没有确定性假设，结果重现了“匹配的内部热力学”以及此处提到的“匹配的内部动力学”或过渡态能量的平缓现象。因此，Albery-Knowles的结果比预期更为普遍。
