Hekman C, Hatefi Y
Department of Molecular and Experimental Medicine, Research Institute of Scripps Clinic, La Jolla, California 92037.
Arch Biochem Biophys. 1991 Jan;284(1):90-7. doi: 10.1016/0003-9861(91)90268-n.
The known subunits of the membrane sector F0 of the bovine mitochondrial ATP synthase complex are subunits b, d, 6, F6, OSCP (oligomycin sensitivity-conferring protein), the DCCD (dicyclohexylcarbodiimide) binding proteolipid, and A6L. The first six subunits were purified from SMP or preparations of the ATP synthase complex, and monospecific antibodies were raised against each. The antisera were shown to be competent for immuno-blotting, and each antiserum recognized a single polypeptide of the expected Mr in preparations of the ATP synthase complex. Immunoblots utilizing antibodies to OSCP and subunits d and 6, which exhibit the same Mr on dodecyl sulfate-polyacrylamide gels, showed clearly that these polypeptides are immunologically distinct. Immunological cross-reactivity was demonstrated between bovine, human, rat, Saccharomyces cerevisiae, Paracoccus denitrificans, and Escherichia coli for subunit 6; between bovine, human, and rat for subunits b, d, OSCP, and F6; and between bovine and rat for the DCCD binding proteolipid. Anti-subunit 6 antiserum, before or after immunopurification against the ATP synthase complex, recognized a single polypeptide in the bovine ATP synthase complex and S. cerevisiae mitochondria, but two polypeptides of different Mr in bovine SMP, human, and rat mitochondria, and Paracoccus and E. coli membranes.
牛线粒体ATP合酶复合体膜部F0已知的亚基有亚基b、d、6、F6、寡霉素敏感性赋予蛋白（OSCP）、二环己基碳二亚胺（DCCD）结合的蛋白脂质以及A6L。前六个亚基是从亚线粒体颗粒（SMP）或ATP合酶复合体的制剂中纯化得到的，并且针对每个亚基制备了单特异性抗体。这些抗血清被证明可用于免疫印迹，并且每种抗血清在ATP合酶复合体的制剂中识别出预期相对分子质量（Mr）的单一多肽。利用针对OSCP以及亚基d和6的抗体进行的免疫印迹（这些亚基在十二烷基硫酸钠-聚丙烯酰胺凝胶上显示相同的Mr）清楚地表明，这些多肽在免疫上是不同的。已证明牛、人、大鼠、酿酒酵母、反硝化副球菌和大肠杆菌的亚基6之间存在免疫交叉反应；牛、人和大鼠的亚基b、d、OSCP和F6之间存在免疫交叉反应；牛和大鼠的DCCD结合蛋白脂质之间存在免疫交叉反应。抗亚基6抗血清在针对ATP合酶复合体进行免疫纯化之前或之后，在牛ATP合酶复合体和酿酒酵母线粒体中识别出单一多肽，但在牛SMP、人及大鼠线粒体以及反硝化副球菌和大肠杆菌膜中识别出两种不同Mr的多肽。
