Identification of mitochondrial gene products by DNA-directed protein synthesis in vitro.
1. A cell-free system, derived from Escherichia coli is highly active in the linked transcription-translation of yeast mtDNA from both wild-type and petite strains. 2. The products of synthesis are short (Mr less than 10 000) hydrophobic polypeptides, which show a high tendency to aggregate in a specific fashion with E. coli and mitochondrial proteins. Aggregation is extremely persistent: alkali, sodium dodecyl sulphate/urea, guanidinium . HCl and carboxymethylation reduce it, but do not eliminate it completely. 3. Nevertheless, results of indirect immunoprecipitation tests suggest that antigenic determinants of cytochrome c oxidase are among the products synthesized. The immunoprecipitation appears specific by criteria including competition experiments and its absence when mtDNA from low complexity petites, retaining only the gene for 21 S rRNA and some flanking sequences, is used to programme protein synthesis. Electrophoretic analysis of material precipitated by anti-cytochrome c oxidase sera reveals four discrete polypeptides with molecular weights of 7400, 6400, 5000 and 4100, which probably represent polypeptide fragments carrying antigenic determinants of cytochrome c oxidase.
D001426:Bacterial Proteins / Q000096:biosynthesis*; D002474:Cell-Free System; D004272:DNA, Mitochondrial / Q000378:metabolism*; D003576:Electron Transport Complex IV / Q000096:biosynthesis / Q000276:immunology; D004926:Escherichia coli / Q000378:metabolism; D008715:Methionine / Q000378:metabolism; D014176:Protein Biosynthesis; D012441:Saccharomyces cerevisiae; D014158:Transcription, Genetic; D014544:Uridine Triphosphate / Q000378:metabolism
D001426:Bacterial Proteins; D004272:DNA, Mitochondrial; D008715:Methionine; D003576:Electron Transport Complex IV; D014544:Uridine Triphosphate