期刊名称：Proceedings of the National Academy of Sciences
印刷版ISSN：0027-8424
电子版ISSN：1091-6490
出版年度：2015
卷号：112
期号：18
页码：5685-5690
DOI：10.1073/pnas.1424353112
语种：English
出版社：The National Academy of Sciences of the United States of America
摘要：SignificanceRespiratory complex I is the largest membrane protein complex in mitochondria and has a central function in energy metabolism. Numerous human diseases are linked with complex I dysfunction or assembly defects. The concerted assembly of more than 40 subunits and the insertion of cofactors is aided by specific chaperones. In addition to eight FeS clusters, complex I comprises a Zn-binding site of unknown function. Combining X-ray structural analysis of complex I crystals with quantum chemical modeling and proteomic and spectroscopic analysis of a purified assembly intermediate, we show that accessory subunit NUMM (human ortholog NDUFS6) binds Zn at the interface of two functional modules of the enzyme complex and is required for a specific step of complex I biogenesis. Mitochondrial proton-pumping NADH:ubiquinone oxidoreductase (respiratory complex I) comprises more than 40 polypeptides and contains eight canonical FeS clusters. The integration of subunits and insertion of cofactors into the nascent complex is a complicated multistep process that is aided by assembly factors. We show that the accessory NUMM subunit of complex I (human NDUFS6) harbors a Zn-binding site and resolve its position by X-ray crystallography. Chromosomal deletion of the NUMM gene or mutation of Zn-binding residues blocked a late step of complex I assembly. An accumulating assembly intermediate lacked accessory subunit N7BM (NDUFA12), whereas a paralog of this subunit, the assembly factor N7BML (NDUFAF2), was found firmly bound instead. EPR spectroscopic analysis and metal content determination after chromatographic purification of the assembly intermediate showed that NUMM is required for insertion or stabilization of FeS cluster N4.
关键词：assembly ; metal protein ; FeS cluster ; NDUFAF2 ; NDUFA12
