A Secondary beta Deuterium Kinetic Isotope Effect in the Chorismate Synthase Reaction.
A Secondary beta Deuterium Kinetic Isotope Effect in the Chorismate Synthase Reaction.
复制标题
分支酸合酶反应中的次级 β 氘动力学同位素效应。
DOI：
--
复制
发表时间：
2000
期刊：
影响因子：
--
通讯作者：
Abell
中科院分区：
文献类型：
--
作者：
Bornemann;Theoclitou;Brune;Webb;Thorneley;Abell
氯酸合成酶(EC 4.6.1.4)是莽草酸途径酶，催化5-烯丙酮基-莽草酸-3-磷酸(EPSP)转化为分支酸。这种酶反应是不寻常的，因为它涉及C-3磷酸和C-6丙酸氢的反式-1，4消除，并且它对还原黄素有绝对要求。已经提出了几种机理来解释该反应的辅因子要求和立体化学，包括自由基机理。本文报道了[4-(2)H]EPSP的合成及其与粗链霉菌和大肠杆菌分支酸合成酶的动力学同位素效应的观察。粗枝杆菌酶在单周转条件下对磷释放的影响为(D)(V)=1.08+/-0.01，对大肠杆菌酶在单周转条件下磷释放的影响为1.10+/-0.02。这种影响最合理的解释是大量的次级贝塔同位素效应。最有可能的是C(3)-O键在非协调的E1键或自由基反应机制中首先断裂。虽然这项研究本身不能排除协调的E2型机制，但在这种机制的过渡状态下，C(3)-O键必须比Pror C(6)-H键的断裂程度要大得多。重要的是，尽管E.Coli和N.crassa酶有不同的限速步骤，但它们的催化机制很可能在化学上是相同的。版权所有2000年学术出版社。
Chorismate synthase (EC 4.6.1.4) is the shikimate pathway enzyme that catalyzes the conversion of 5-enolpyruvylshikimate 3-phosphate (EPSP) to chorismate. The enzyme reaction is unusual because it involves a trans-1,4 elimination of the C-3 phosphate and the C-6 proR hydrogen and it has an absolute requirement for reduced flavin. Several mechanisms have been proposed to account for the cofactor requirement and stereochemistry of the reaction, including a radical mechanism. This paper describes the synthesis of [4-(2)H]EPSP and the observation of kinetic isotope effects using this substrate with both Neurospora crassa and Escherichia coli chorismate synthases. The magnitude of the effects were (D)(V) = 1.08 +/- 0.01 for the N. crassa enzyme and 1.10 +/- 0.02 on phosphate release under single-turnover conditions for the E. coli enzyme. The effects are best rationalised as substantial secondary beta isotope effects. It is most likely that the C(3)-O bond is cleaved first in a nonconcerted E1 or radical reaction mechanism. Although this study alone cannot rule out a concerted E2-type mechanism, the C(3)-O bond would have to be substantially more broken than the proR C(6)-H bond in a transition state of such a mechanism. Importantly, although the E. coli and N. crassa enzymes have different rate limiting steps, their catalytic mechanisms are most likely to be chemically identical. Copyright 2000 Academic Press.
霍乱弧菌神经氨酸酶催化的动力学同位素效应研究。
DOI：
10.1042/bj2940653
发表时间：
1993
期刊：
The Biochemical journal
影响因子：
--
作者：
Guo,X;Sinnott,ML
通讯作者：
Sinnott,ML
底物类似物作为粗糙脉孢菌双功能分支酸合酶的机械探针。
影响因子：
2.9
作者：
Lauhon,CT;Bartlett,PA
通讯作者：
Bartlett,PA