Catalysis of acetoin formation by brewers' yeast pyruvate decarboxylase isozymes.
Catalysis of acetoin formation by brewers' yeast pyruvate decarboxylase isozymes.
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啤酒酵母丙酮酸脱羧酶同工酶催化乙偶姻形成。
DOI：
10.1021/bi00212a012
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发表时间：
1993
期刊：
影响因子：
2.9
通讯作者：
Washabaugh,MW
中科院分区：
文献类型：
--
作者：
Stivers,JT;Washabaugh,MW
H、D 或 T) 形成乙偶姻以及主要动力学同位素对反应的影响。 PDC 同工酶混合物和 «4 同工酶 («4-PDC) 在存在和不存在非底物变构效应物丙酮酰胺的情况下对乙偶姻形成具有不同的稳态动力学参数和同位素效应：丙酮酰胺活化是通过乙醛/PDC 三元复合物的稳定而发生的。主要 L (V/K) 型（L= D 或 T）同位素对从 «4-PDC 结合的 HETDP 转移的 C (a)-质子的影响程度没有提供 Swain-Schaad 关系显着破坏的证据，而这表明假定的 C (a)-碳负离子/烯胺中间体在 HETDP 和产物之间的分配。氘初级动力学同位素效应的底物浓度依赖性为从 4-PDC 结合的 HETDP 转移 C (a)-质子的内在同位素效应 4.1 提供了证据。乙偶姻形成中对 [1, 2-14C] 乙醛的 1.10±0.02 倍 14C 同位素歧视与逐步机制不一致，其中添加步骤发生在作为离散酶结合中间体的 C (a)-碳负离子/烯胺的限速形成之后，并为过渡态碳-碳键形成的重要组成部分的协同反应机制提供了证据。丙酮酸脱羧酶(PDC) 1（2-含氧酸羧基裂解酶；EC 4.1. 1.1）是硫胺素二磷酸 (TDP, 1) 依赖性酶，可催化丙酮酸不可逆非氧化脱羧形成乙醛（方案 I，上途径）（Alvarez 等人，1991）。 PDC 还催化两个乙醛分子之间的醛醇型缩合反应，形成酮醇乙偶姻 (2)（方案 I；下游途径）（Chen & Jordan，1984）。由 2-(1-羟乙基) 硫胺素二磷酸 (HETDP, 4) 衍生的 C(a)-碳负离子/烯胺 (3) 参与了几种 TDP 依赖性酶催化的 HETDP 和羰基化合物之间的醇醛型加成反应 (Kluger, 1992)。共轭 C (a)-烯胺已被观察为 PDC 催化脱羧和其他涉及苯环上具有强吸电子取代基的 (£)-2-oxo-4-(4-R-苯基)-3-丁烯酸的反应过程中的中间体 (Zeng et al., 1991)。
H, D, or T) to form acetoin and the primary kinetic isotope effects on the reaction. The PDC isozyme mixture and «4 isozyme («4-PDC) have different steady-state kinetic parameters and isotope effects for acetoin formation in the presence and absence of the nonsubstrate allosteric effector pyruvamide: pyruvamideactivation occurs by stabilization of the acetaldehyde/PDC ternary complex. The magnitudes of primary L (V/K)-type (L= D or T) isotope effects on C (a)-proton transfer from «4-PDC-bound HETDP provide no evidence for significant breakdown of the Swain-Schaad relationship that would indicate partitioning of the putative C (a)-carbanion/enamine intermediate between HETDP and products. The substrate concentration dependence of the deuterium primary kinetic isotope effects providesevidence for an intrinsic isotope effect of 4.1 for C (a)-proton transfer from «4-PDC-bound HETDP. A 1.10±0.02-fold 14C isotope discrimination against [1, 2-14C] acetaldehyde in acetoin formation is inconsistent with a stepwise mechanism, in which the addition step occurs after rate-limiting formation of the C (a)-carbanion/enamine as a discrete enzymebound intermediate, and provides evidence for a concerted reaction mechanism with an important component of carbon-carbon bond formation in the transition state.Pyruvate decarboxylase (PDC) 1 (2-oxo-acid carboxy-lyase; EC 4.1. 1.1) is a thiamin diphosphate (TDP, 1) dependent enzyme that catalyzes the irreversible nonoxidative decar-boxylation of pyruvate to form acetaldehyde (Scheme I, upper pathway)(Alvarez etal., 1991). PDC also catalyzes an aldoltype condensation reaction between two molecules of ace-taldehyde to form the-ketol acetoin (2)(Scheme I; lower pathway)(Chen & Jordan, 1984). The C (a)-carbanion/enamine (3) derived from 2-(1-hydroxyethyl) thiamin diphosphate (HETDP, 4) has been implicated in aldol-type addition reactions between HETDP and carbonyl compounds catalyzed by several TDP-dependent enzymes (Kluger, 1992). Conjugated C (a)-enamines have been observed as intermediates during catalysis by PDC of decarboxylation and other reactions involving (£)-2-oxo-4-(4-R-phenyl)-3-butenoic acids that have strong electron-withdrawing substituents on the phenyl ring (Zeng et al., 1991).