Beattie D S, Clejan L
J Bioenerg Biomembr. 1980 Apr;12(1-2):35-45. doi: 10.1007/BF00745011.
Mitochondrial translation produces obtained from yeast cells labeled in vivo in the presence of cycloheximide were separated by dodecylsulfate polyacrylamide gel electrophoresis. The labeled band, with a molecular weight of 30,000 corresponding to cytochrome b, was excised and subsequently transferred to a second gel. After electrophoretic separation, two labeled polypetides with apparent molecular weights of 67,000 and 27,000 became visible in addition to the cytochrome b band of 30,000 molecular weight. Heating of the cytochrome b band prior to transfer resulted in an increase in the amount of the labeled polypeptides migrating with a molecular weight of 67,000. Longer exposure during autoradiography of the gels of mitochondrial translation products resulted in the appearance of a double band with an apparent molecular weight of 67,000. Limited proteolysis of this 67,000 dalton protein with Staphylococcus aureus V8 protease revealed a peptide map similar to that obtained after proteolysis of cytochrome b. These results suggest that the polypeptide with an apparent molecular weight of 67,000 represents an aggregate of cytochrome b that is either present as such in the membrane or is formed in vitro during the experimental manipulation to prepare mitochondria for gel electrophoresis.
在环己酰亚胺存在的情况下，对体内标记的酵母细胞所产生的线粒体翻译产物进行十二烷基硫酸钠聚丙烯酰胺凝胶电泳分离。切除分子量为30,000且对应于细胞色素b的标记条带，随后将其转移至另一凝胶上。电泳分离后，除了分子量为30,000的细胞色素b条带外，还可见两条表观分子量分别为67,000和27,000的标记多肽。转移前对细胞色素b条带进行加热，导致分子量为67,000的迁移标记多肽量增加。对线粒体翻译产物凝胶进行放射自显影时延长曝光时间，出现了一条表观分子量为67,000的双链带。用金黄色葡萄球菌V8蛋白酶对这条67,000道尔顿的蛋白质进行有限的蛋白酶解，显示出与细胞色素b蛋白酶解后所得相似的肽图。这些结果表明，表观分子量为67,000的多肽代表细胞色素b的聚集体，其要么以这种形式存在于膜中，要么在为凝胶电泳制备线粒体的实验操作过程中在体外形成。
