Molecular mechanism of regulation of the pyruvate dehydrogenase complex from E-coli
Molecular mechanism of regulation of the pyruvate dehydrogenase complex from E-coli
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DOI：
10.1021/bi971845z
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发表时间：
1997-12-16
期刊：
影响因子：
2.9
通讯作者：
Hübner, G
中科院分区：
文献类型：
--
作者：
Hennig, J;Kern, G;Hübner, G
来自大肠杆菌的丙酮酸脱氢酶多酶复合物显示出反应速率对底物浓度的S形依赖性，当在生理浓度的辅因子二磷酸硫胺素存在下形成产物时。为了阐明这种调节的分子机制，底物丙酮酸对辅酶-蛋白质相互作用的影响已经使用几种辅酶类似物进行了研究。在底物丙酮酸的存在下，所有具有辅酶活性的类似物的观察到的结合常数都增加，而在丙酮酸的存在下，所有无辅酶活性的类似物的观察到的结合常数都没有改变。这表明反应-中间体-辅酶复合物与蛋白质的结合亲和力增加。由于辅因子结合和解离在生理浓度的硫胺二磷酸是缓慢的催化反应相比，缓慢过渡到活性状态的酶发生。在降低丙酮酸浓度之后！观察到相反的效果，即二磷酸硫胺素从酶上解离。辅因子结合的这种缓慢的底物依赖性增强使得丙酮酸脱氢酶复合物能够被其底物丙酮酸有效调节。
The pyruvate dehydrogenase multienzyme complex from E, coli shows a sigmoidal dependency of the reaction rate on the substrate concentration when product formation is followed in the presence of physiological concentrations of the cofactor thiamin diphosphate. To elucidate the molecular mechanism of this regulation, the influence of the substrate pyruvate on the coenzyme-protein interaction has been investigated using several coenzyme analogues. The observed binding constants of all coenzymatically active analogues are increased in the presence of the substrate pyruvate, whereas those of all coenzymatically inactive analogues are not altered in the presence of pyruvate. This points to an increased binding affinity of a reaction-intermediate-coenzyme complex to the protein. Since cofactor binding and dissociation at physiological concentrations of thiamin diphosphate are slow compared to the catalytic reaction, a slow transition to the active state of the enzyme occurs. After lowering the pyruvate concentration! the opposite effect, a dissociation of the thiamin diphosphate from the enzyme is observed. This slow substrate dependent enhancement of cofactor binding enables efficient regulation of the pyruvate dehydrogenase complex by its substrate pyruvate.