Landry Aaron P, Cheng Zishuo, Ding Huangen
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Department of Biological Sciences, Louisiana State University, Baton Rouge, LA 70803, USA.
Free Radic Biol Med. 2015 Apr;81:119-27. doi: 10.1016/j.freeradbiomed.2015.01.017. Epub 2015 Jan 30.
The human mitochondrial outer membrane protein mitoNEET is a newly discovered target of the type 2 diabetes drug pioglitazone. Structurally, mitoNEET is a homodimer with each monomer containing an N-terminal transmembrane α helix tethered to the mitochondrial outer membrane and a C-terminal cytosolic domain hosting a redox-active [2Fe-2S] cluster. Genetic studies have shown that mitoNEET has a central role in regulating energy metabolism in mitochondria. However, the specific function of mitoNEET remains largely elusive. Here we find that the mitoNEET [2Fe-2S] clusters can be efficiently reduced by Escherichia coli thioredoxin reductase and glutathione reductase in an NADPH-dependent reaction. Purified human glutathione reductase has the same activity as E. coli thioredoxin reductase and glutathione reductase to reduce the mitoNEET [2Fe-2S] clusters. However, rat thioredoxin reductase, a human thioredoxin reductase homolog that contains selenocysteine in the catalytic center, has very little or no activity to reduce the mitoNEET [2Fe-2S] clusters. N-ethylmaleimide, a potent thiol modifier, completely inhibits human glutathione reductase from reducing the mitoNEET [2Fe-2S] clusters, indicating that the redox-active disulfide in the catalytic center of human glutathione reductase may be directly involved in reducing the mitoNEET [2Fe-2S] clusters. Additional studies reveal that the reduced mitoNEET [2Fe-2S] clusters in mouse heart cell extracts can be reversibly oxidized by hydrogen peroxide without disruption of the clusters, suggesting that the mitoNEET [2Fe-2S] clusters may undergo redox transition to regulate energy metabolism in mitochondria in response to oxidative signals.
人类线粒体外膜蛋白米托萘醌是2型糖尿病药物吡格列酮新发现的靶点。从结构上看，米托萘醌是一种同型二聚体，每个单体包含一个与线粒体外膜相连的N端跨膜α螺旋和一个含有氧化还原活性[2Fe-2S]簇的C端胞质结构域。遗传学研究表明，米托萘醌在调节线粒体能量代谢中起核心作用。然而，米托萘醌的具体功能仍 largely难以捉摸。在这里，我们发现米托萘醌的[2Fe-2S]簇可以在NADPH依赖的反应中被大肠杆菌硫氧还蛋白还原酶和谷胱甘肽还原酶有效还原。纯化的人谷胱甘肽还原酶与大肠杆菌硫氧还蛋白还原酶和谷胱甘肽还原酶具有相同的活性来还原米托萘醌的[2Fe-2S]簇。然而，大鼠硫氧还蛋白还原酶，一种在催化中心含有硒代半胱氨酸的人硫氧还蛋白还原酶同源物，对还原米托萘醌的[2Fe-2S]簇几乎没有或没有活性。N-乙基马来酰亚胺，一种有效的硫醇修饰剂，完全抑制人谷胱甘肽还原酶还原米托萘醌的[2Fe-2S]簇，表明人谷胱甘肽还原酶催化中心的氧化还原活性二硫键可能直接参与还原米托萘醌的[2Fe-2S]簇。进一步的研究表明，小鼠心脏细胞提取物中还原的米托萘醌[2Fe-2S]簇可以被过氧化氢可逆氧化而不破坏簇，这表明米托萘醌的[2Fe-2S]簇可能经历氧化还原转变以响应氧化信号来调节线粒体中的能量代谢。
