Vázquez-Acevedo M, Antaramian A, Corona N, González-Halphen D
Departamento de Bioenergética, Universidad Nacional Autónoma de México, México, D.F.
J Bioenerg Biomembr. 1993 Aug;25(4):401-10. doi: 10.1007/BF00762466.
The existence of tissue-specific isozymes of cytochrome c oxidase has been widely documented. We have now studied if there are differences between subunits of mitochondrial bc1 complexes isolated from liver and heart. For this purpose, we have developed a method for the purification of an active ubiquinol-cytochrome c oxidoreductase from adult bovine liver that includes solubilization of submitochondrial particles with deoxycholate, ammonium acetate fractionation, resolubilization with dodecyl maltoside, and ion exchange chromatography. The electrophoretic pattern of the liver preparation showed the presence of 11 subunits, with apparent molecular weights identical to the ones reported for the heart complex. Western blot analysis and isoelectric focusing followed by two-dimensional gels of bc1 complexes from liver and heart were compared, and no qualitative differences were observed. In addition, the high-molecular-weight subunits of the purified complexes from both tissues, subunits I, II, V, and VI, were isolated by PAGE in the presence of Coomasie Blue and subjected to limited proteolysis and to chemical digestion with cyanogen bromide and BNPS-skatol, and the peptide patterns were compared. Finally, two of the small-molecular-weight subunits from the liver complex were isolated (subunits VII and X), partially analyzed by amino terminal sequencing, and found to be identical with the reported sequence of their heart counterparts. The data suggest that, in contrast to the case of cytochrome c oxidase, bc1 complexes from liver and heart do not exhibit tissue-specific differences.
细胞色素c氧化酶组织特异性同工酶的存在已被广泛记载。我们现在研究了从肝脏和心脏分离出的线粒体bc1复合物亚基之间是否存在差异。为此，我们开发了一种从成年牛肝脏中纯化活性泛醇-细胞色素c氧化还原酶的方法，该方法包括用脱氧胆酸盐溶解亚线粒体颗粒、醋酸铵分级分离、用十二烷基麦芽糖苷再溶解以及离子交换色谱法。肝脏制剂的电泳图谱显示存在11个亚基，其表观分子量与报道的心脏复合物的分子量相同。比较了肝脏和心脏bc1复合物的蛋白质印迹分析以及等电聚焦后二维凝胶的结果，未观察到定性差异。此外，通过在考马斯亮蓝存在下的聚丙烯酰胺凝胶电泳分离了来自两种组织的纯化复合物的高分子量亚基，即亚基I、II、V和VI，并对其进行了有限的蛋白水解以及用溴化氰和BNPS-斯卡特ol进行化学消化，然后比较了肽图谱。最后，分离出了肝脏复合物的两个小分子亚基（亚基VII和X），通过氨基末端测序进行了部分分析，发现它们与报道的心脏对应亚基序列相同。数据表明，与细胞色素c氧化酶的情况不同，肝脏和心脏的bc1复合物没有表现出组织特异性差异。
