Modulation of mitochondrial complex I activity by reversible Ca2+ and NADH mediated superoxide anion dependent inhibition.
Modulation of mitochondrial complex I activity by reversible Ca2+ and NADH mediated superoxide anion dependent inhibition.
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DOI：
10.1021/bi049803f
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发表时间：
2004-07
期刊：
影响因子：
2.9
通讯作者：
Hesham A. Sadek;P. Szweda;L. Szweda
中科院分区：
文献类型：
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作者：
Hesham A. Sadek;P. Szweda;L. Szweda
复合物I是线粒体呼吸链的关键组分，由于心脏缺血/再灌注而表现出活性降低。心肌缺血/再灌注与线粒体Ca（2+）和促氧化剂水平的增加有关。在目前的体外研究中，我们寻找证据的机制之间的联系Ca（2+），促氧化剂，并利用线粒体从大鼠心脏中分离的复合物I的抑制。我们的研究结果表明，另外的Ca（2+）的溶解线粒体的结果在复合物I活性的损失。Ca（2+）在较窄的生理相关pH范围内，在低微摩尔浓度下诱导复合物I活性最大降低约35%。活性损失需要还原当量形式的NADH，并且在加入EGTA后不会逆转。抗氧化剂N-乙酰半胱氨酸和超氧化物歧化酶，但不是过氧化氢酶，防止抑制，表明超氧阴离子（O2（*-））的参与失活过程。重要的是，巯基还原剂DTT能够完全恢复复合物I的活性，这意味着在失活过程中形成次磺酸和/或半胱氨酸的二硫化物衍生物。最后，如果存在NADH，复合物I可以在Ca（2+）去除后内源性地重新激活，并且允许酶催化周转。因此，本研究提供了已知在心脏缺血/再灌注期间发生的三种改变之间的机制联系，即线粒体Ca（2+）积累、自由基产生和复合物I抑制。这些过程的可逆性表明Ca（2+）处理的氧化还原调节。
Complex I, a key component of the mitochondrial respiratory chain, exhibits diminished activity as a result of cardiac ischemia/reperfusion. Cardiac ischemia/reperfusion is associated with increases in the levels of mitochondrial Ca(2+) and pro-oxidants. In the current in vitro study, we sought evidence for a mechanistic link between Ca(2+), pro-oxidants, and inhibition of complex I utilizing mitochondria isolated from rat heart. Our results indicate that addition of Ca(2+) to solubilized mitochondria results in loss in complex I activity. Ca(2+) induced a maximum decrease in complex I activity of approximately 35% at low micromolar concentrations over a narrow physiologically relevant pH range. Loss in activity required reducing equivalents in the form of NADH and was not reversed upon addition of EGTA. The antioxidants N-acetylcysteine and superoxide dismutase, but not catalase, prevented inhibition, indicating the involvement of superoxide anion (O2(*-)) in the inactivation process. Importantly, the sulfhydryl reducing agent DTT was capable of fully restoring complex I activity implicating the formation of sulfenic acid and/or disulfide derivatives of cysteine in the inactivation process. Finally, complex I can reactivate endogenously upon Ca(2+) removal if NADH is present and the enzyme is allowed to turnover catalytically. Thus, the present study provides a mechanistic link between three alterations known to occur during cardiac ischemia/reperfusion, mitochondrial Ca(2+) accumulation, free radical production, and complex I inhibition. The reversibility of these processes suggests redox regulation of Ca(2+) handling.