Goodman S I, Axtell K M, Bindoff L A, Beard S E, Gill R E, Frerman F E
Department of Pediatrics, University of Colorado School of Medicine, Denver 80262.
Eur J Biochem. 1994 Jan 15;219(1-2):277-86. doi: 10.1111/j.1432-1033.1994.tb19939.x.
Electron-transfer flavoprotein-ubiquinone oxidoreductase (ETF-QO) in the inner mitochondrial membrane accepts electrons from electron-transfer flavoprotein which is located in the mitochondrial matrix and reduces ubiquinone in the mitochondrial membrane. The two redox centers in the protein, FAD and a [4Fe4S]+2,+1 cluster, are present in a 64-kDa monomer. We cloned several cDNA sequences encoding the majority of porcine ETF-QO and used these as probes to clone a full-length human ETF-QO cDNA. The deduced human ETF-QO sequence predicts a protein containing 617 amino acids (67 kDa), two domains associated with the binding of the AMP moiety of the FAD prosthetic group, two membrane helices and a motif containing four cysteine residues that is frequently associated with the liganding of ferredoxin-like iron-sulfur clusters. A cleavable 33-amino-acid sequence is also predicted at the amino terminus of the 67-kDa protein which targets the protein to mitochondria. In vitro transcription and translation yielded a 67-kDa immunoprecipitable product as predicted from the open reading frame of the cDNA. The human cDNA was expressed in Saccharomyces cerevisiae, which does not normally synthesize the protein. The ETF-QO is synthesized as a 67-kDa precursor which is targeted to mitochondria and processed in a single step to a 64-kDa mature form located in the mitochondrial membrane. The detergent-solubilized protein transfers electrons from ETF to the ubiquinone homolog, Q1, indicating that both the FAD and iron-sulfur cluster are properly inserted into the heterologously expressed protein.
线粒体内膜中的电子传递黄素蛋白 - 泛醌氧化还原酶（ETF - QO）从位于线粒体基质中的电子传递黄素蛋白接受电子，并还原线粒体内膜中的泛醌。该蛋白质中的两个氧化还原中心，即黄素腺嘌呤二核苷酸（FAD）和一个[4Fe4S]+2，+1簇，存在于一个64 kDa的单体中。我们克隆了几个编码猪ETF - QO大部分序列的cDNA序列，并将这些序列用作探针来克隆全长人ETF - QO cDNA。推导的人ETF - QO序列预测该蛋白质含有617个氨基酸（67 kDa），有两个与FAD辅基的AMP部分结合相关的结构域，两个膜螺旋以及一个包含四个半胱氨酸残基的基序，该基序通常与铁氧化还原蛋白样铁硫簇的配位有关。在67 kDa蛋白质的氨基末端还预测有一个可切割的33个氨基酸的序列，该序列将蛋白质靶向线粒体。体外转录和翻译产生了一个67 kDa的可免疫沉淀产物，这与cDNA的开放阅读框预测的一致。人cDNA在通常不合成该蛋白质的酿酒酵母中表达。ETF - QO以67 kDa的前体形式合成，该前体靶向线粒体并一步加工成位于线粒体内膜中的64 kDa成熟形式。去污剂溶解的蛋白质将电子从ETF转移到泛醌同系物Q1，表明FAD和铁硫簇都正确插入到异源表达的蛋白质中。
