CATALYTIC SIGNIFICANCE OF BINARY ENZYME-ALDEHYDE COMPLEXES IN THE LIVER ALCOHOL-DEHYDROGENASE REACTION
CATALYTIC SIGNIFICANCE OF BINARY ENZYME-ALDEHYDE COMPLEXES IN THE LIVER ALCOHOL-DEHYDROGENASE REACTION
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DOI：
10.1111/j.1432-1033.1984.tb08036.x
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发表时间：
1984-01-01
期刊：
影响因子：
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通讯作者：
PETTERSSON, G
中科院分区：
文献类型：
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作者：
ANDERSSON, P;KVASSMAN, J;PETTERSSON, G
[马]肝醇脱氢酶与NADH和醛底物的相互作用的特征在于相对于三元复合物的形成由明显的非优选途径，其中涉及中间形成的二元酶.醛复合物。报道了二甲基氨基肉桂醛（DACA）与游离酶结合和NADH与酶·DACA复合物结合的速率常数估计值。NADH（或NAD）与肝醇脱氢酶结合的速率不受DACA与酶结合的可检测影响，但NADH解离速率降低约1/3。6. NADH诱导的酶对DACA的亲和力增加类似地归因于解离速率降低而不是醛的缔合速率增加。DACA比辅酶更快地从酶·醛复合物中解离，并且在其与游离酶的相互作用中显示出比NADH更高的缔合速率常数。典型醛底物的酶促还原可能符合速率方程，该速率方程在实验上与辅酶结合先于底物结合的强制顺序机制的速率方程难以区分，并且该速率方程将获得而不管实际上优选哪种三元复合物形成途径。速率方程没有提供可靠的信息，在三元络合物系统中的配体结合的顺序。流动分析表明，辅酶和底物实际上是结合在随机顺序的肝醇脱氢酶在酶还原醛的NADH。用于形成三元复合物的酶-醛途径是完全动力学活性的，并且当醛浓度超过游离酶半饱和所需的那些时，经由该途径的反应流可能占主导地位。二元酶·醛复合物对于酶的速率行为似乎是不重要的，但可以对催化反应流提供最显著的甚至是主要的贡献。
The interaction of [horse] liver alcohol dehydrogenase with NADH and aldehyde substrates was characterized with respect to ternary-complex formation by the apparently non-preferred pathway which involves intermediate formation of binary enzyme.cntdot.aldehyde complexes. Rate constant estimates are reported for dimethylaminocinnamaldehyde (DACA) binding to free enzyme and for NADH binding to the enzyme.cntdot.DACA complex. The rate of NADH (or NAD) association to liver alcohol dehydrogenase is not detectably affected by DACA binding to the enzyme, but the NADH dissociation rate decreases by a factor of .apprx. 6. The NADH-induced increase in affinity of the enzyme for DACA is similarly attributable to a decreased dissociation rate rather than an increased association rate of the aldehyde. DACA dissociates much more rapidly than coenzyme from the enzyme.cntdot.NADH.cntdot.aldehyde complex and shows a higher association rate constant than NADH in its interaction with free enzyme. The enzymic reduction of typical aldehyde substrates will probably conform to a rate equation which is experimentally indistinguishable from that of a compulsory-order mechanism with coenzyme binding preceding substrate binding, and this rate equation will obtain irrespective of which pathway for ternary-complex formation is actually preferred. Rate equations provide no reliable information about the order of ligand binding in ternary-complex systems. A flow analysis is presented which indicates that coenzyme and substrate are actually bound in random order to liver alcohol dehydrogenase during the enzyme reduction of aldehydes by NADH. The enzyme.cntdot.aldehyde pathway for ternary-complex formation is fully kinetically competent, and reaction flow via this pathway may predominate when aldehyde concentrations exceed those required for half-saturation of free enzyme. Binary enzyme.cntdot.aldehyde complexes are seemingly insignificant with respect to the rate behavior of the enzyme, but may provide most significant and even predominant contributions to the catalytic reaction flow.