Proteins coded by mitochondrial DNA of mammalian cells.

Abstract

Improved conditions are described for the maintenance of protein synthesis by isolated liver mitochondria. The products of synthesis in vitro are first attached to a 55-S mitoribosome (mitochondrial-ribosome) but move off the mitoribosome as completed peptide chains in less than 15 min at 30 °C. Over a 1-h period of incubation there is no change in the spectrum of molecular weights of the peptide being synthesized. The protein synthesized by isolated hamster-liver mitochondria were compared with those synthesized by whole cells under conditions where cytoplasmic protein synthesis had been blocked by emetine. The proteins synthesized by whole cell mitochondria were indistinguishable from those synthesized by isolated mitochondria. By the combined use of emetine, chlorampheinicol and ethidium bromide as specific inhibitors of cytoribosomal and mitoribosomal protein synthesis it was possible to calculate that 7% of the total mitochondrial protein was coded by the mitochondrial genome. A phosphate buffer at pH 11.5 solubilized about 90% of the total protein of the mitochondrion but left over 90% of the protein coded by the mitochondrial genome as an insoluble residue. This fraction on polyacrylamide in the presence of sodium dodecylsuphate and mercaptoethanol was shown to contain at least 20 distinct proteins. At least 10 of these were coded by the mitochondrial genome and their molecular weights ranged from 14000 to 50000. These proteins together with the mitochondrial tRNA and the mitoribosomal RNA account for most of the information available in the mitochondrial genome.