Qualitative and comparative nature of mitochondrial translation products in mammalian cells.
A method has been described for the efficient incorporation of [35S]methionine into isolated mitochondrial particles from various mammalian tissues. The method involves the incubation of digitonin-treated mitochondrial particles (mitoplasts) in a low sucrose medium. Electrophoretic analysis of 35S-labeled products on sodium dodecyl sulfate-polyacrylamide gels under reducing conditions shows that mitoplasts from Ehrlich ascites cells, mouse liver, and rat liver synthesize 19-24 polypeptide species including some high molecular weight components in the size range of 1.0 X 10(5). The polypeptide species synthesized in the mitoplast system resemble the cycloheximide-resistant products synthesized in the intact cells with respect to size distribution and total number, although significant quantitative differences between the two systems are observed. Experiments on pulse--chase analysis of 35S-labeled mitochondrial products and the effects of protease inhibitors on the electrophoretic profiles suggest no significant proteolytic degradation during the incubation or analysis. Further, control experiments with nuclease-treated mitoplasts and use of specific protein synthesis inhibitors show that all of the labeled polypeptides are the intramitochondrial translation products. Extensive comparison between the products synthesized in Ehrlich ascites and mouse and rat liver mitochondria, using one- and two-dimensional gels under denaturing conditions, shows striking variations, suggesting possible heterogeneity.
D000818:Animals; D002286:Carcinoma, Ehrlich Tumor / Q000378:metabolism; D002478:Cells, Cultured; D004272:DNA, Mitochondrial / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D066298:In Vitro Techniques; D051379:Mice; D008928:Mitochondria / Q000378:metabolism*; D008930:Mitochondria, Liver / Q000378:metabolism; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000378:metabolism; D000077278:RNA, Mitochondrial; D051381:Rats
D004272:DNA, Mitochondrial; D012333:RNA, Messenger; D000077278:RNA, Mitochondrial; C029305:mitochondrial messenger RNA