Maklashina E O, Sled' V D, Vinogradov A D
Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia 19104.
Biokhimiia. 1994 Jul;59(7):946-57.
Isolated Complex I exists in two forms. The active form catalyzes the rapid rotenone-sensitive, N-ethylmaleimide-insensitive NADH : Q1 reductase reaction. The inactive form is inhibited by N-ethylmaleimide and catalyzes the rotenone-sensitive ubiquinone reduction with a prominent lag phase. The inactive enzyme is transformed into its active form after rapid reduction by NADH and slow (compared with the steady-state turnover number) oxidation by quinone. The rate of activation is strongly temperature-dependent (the activation energy is 170 kJ/mol) and influenced by pH and divalent cations. The active enzyme is quite stable (hours at 0 degrees C) but it is spontaneously deactivated at high temperature (the activation energy is 245 kJ/mol). The active/inactive transition parameters are qualitatively and quantitatively similar for the isolated and membrane-bound Complex I. The extent of the isolated Complex I activation in the presence of NADH depends on the concentration of the added quinone. The concentration of quinone needed for the half-maximal activation is 10 times less than the KQ1m value in the steady-state NADH : Q1 reductase reaction. The data obtained suggest that the free energy of NADH oxidation in the respiratory chain is partly utilized to maintain the catalytically competent Complex I conformation.
分离出的复合体I以两种形式存在。活性形式催化快速的鱼藤酮敏感、N-乙基马来酰亚胺不敏感的NADH：Q1还原酶反应。非活性形式被N-乙基马来酰亚胺抑制，并催化鱼藤酮敏感的泛醌还原，伴有明显的滞后阶段。非活性酶在被NADH快速还原并被醌缓慢氧化（与稳态周转数相比）后转变为活性形式。活化速率强烈依赖温度（活化能为170 kJ/mol），并受pH值和二价阳离子影响。活性酶相当稳定（在0℃下可保持数小时），但在高温下会自发失活（活化能为245 kJ/mol）。分离的和膜结合的复合体I的活性/非活性转变参数在定性和定量上相似。在NADH存在下，分离的复合体I的活化程度取决于添加醌的浓度。半最大活化所需的醌浓度比稳态NADH：Q1还原酶反应中的KQ1m值低10倍。所获得的数据表明，呼吸链中NADH氧化的自由能部分用于维持具有催化活性的复合体I构象。
