Generation of superoxide by the mitochondrial Complex I
Generation of superoxide by the mitochondrial Complex I
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DOI：
10.1016/j.bbabio.2006.03.013
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发表时间：
2006-05-01
影响因子：
4.3
通讯作者：
Vinogradov, Andrei D.
中科院分区：
文献类型：
--
作者：
Grivennikova, Vera G.;Vinogradov, Andrei D.
用琥珀酸或NADH呼吸，测量了由内向外耦合的牛心亚线粒体颗粒产生的超氧化物。鱼藤酮和解偶联剂抑制了琥珀酸氧化产物的生成，表明琥珀酸氧化过程中产生的超氧化物大部分来自络合物I的单价氧还原。高浓度NADH(1 MM)下呼吸过程中超氧化物(O-2(.-))的生成速率显著低于琥珀酸氧化过程。此外，在1mMNADH存在下，琥珀酸支持的O-2(.-)的产生显著减少。滴定曲线，即超氧化物产生的初始速率与NADH浓度的关系，呈钟形，最大速率(50亩NADH)接近琥珀酸根。NAD+和NADH：乙酰基-NAD+转氢酶分别抑制了琥珀酸负载的反应，其表观KI接近其K-m。我们的结论是：(I)在人工实验条件下，呼吸链产生的大部分超氧化物是由络合物I的某些氧化还原成分(很可能是还原或自由基形式的FMN)形成的，(Ii)络合物I中的NADH(F位)和NAD+(R位)有两个不同的结合部位，提供了底物-核苷酸与酶红氧化成分(S)的可及性，F位是NADH氧化的入口，而R位是反向电子转移和单价氧还原；(Iii)在生理条件下(高浓度的NADH和NAD+)，复合体I不太可能负责线粒体超氧化物的产生。我们认为，如果线粒体确实在生理相关的条件下参与了ROS控制的过程，那么线粒体基质中应该存在与NAD(P)H/NAD(P)+对处于平衡状态的特异性NAD(P)H：产生氧超氧化物(过氧化氢)氧化还原酶(S)。(C)2006爱思唯尔B.V.保留所有权利。
Superoxide production by inside-out coupled bovine heart submitochondrial particles, respiring with succinate or NADH, was measured. The succinate-supported production was inhibited by rotenone and uncouplers, showing that most part of superoxide produced during succinate oxidation is originated from univalent oxygen reduction by Complex I. The rate of the superoxide (O-2(.-)) production during respiration at a high concentration of NADH (1 mM) was significantly lower than that with succinate. Moreover, the succinate-supported O-2(.-) production was significantly decreased in the presence of I mM NADH. The titration curves, i.e., initial rates of superoxide production versus NADH concentration, were bell-shaped with the maximal rate (at 50 mu M NADH) approaching that seen with succinate. Both NAD+ and acetyl-NAD+ inhibited the succinate-supported reaction with apparent Ki's close to their K-m's in the Complex I-catalyzed succinate-dependent energy-linked NAD+ reduction (reverse electron transfer) and NADH:acetyl-NAD+ transhydrogenase reaction, respectively. We conclude that: (i) under the artificial experimental conditions the major part of superoxide produced by the respiratory chain is formed by some redox component of Complex I (most likely FMN in its reduced or free radical form); (ii) two different binding sites for NADH (F-site) and NAD+ (R-site) in Complex I provide accessibility of the substrates-nucleotides to the enzyme red-ox component(s); F-site operates as an entry for NADH oxidation, whereas R-site operates in the reverse electron transfer and univalent oxygen reduction; (iii) it is unlikely that under the physiological conditions (high concentrations of NADH and NAD+) Complex I is responsible for the mitochondrial superoxide generation. We propose that the specific NAD(P) H:oxygen superoxide (hydrogen peroxide) producing oxidoreductase(s) poised in equilibrium with NAD(P)H/NAD(P)+ couple should exist in the mitochondrial matrix, if mitochondria are, indeed, participate in ROS-controlled processes under physiologically relevant conditions. (c) 2006 Elsevier B.V. All rights reserved.