Formate as a Specific Label for Mitochondrial Translational Products

Abstract

Abstract Based on the well authenticated observation that the mitochondrial protein-synthesizing system utilizes fMet for chain initiation, we have tested the specificity of radioactive formate as a potential label for such nascent polypeptide chains in Saccharomyces cerevisiae. We find that at short incubation times, protein-bound formate is localized exclusively on mitochondrial—and absent from extramitochondrial—ribosomes, polysomes, and subunits. In the presence of 0.5 mm puromycin it is quantitatively converted to fMet-(peptidyl)-puromycin derivatives; hence the formation of labeled fMet-puromycin itself, extracted from reaction mixtures continuously exposed to the inhibitor, can be utilized as a quantitative measure of mitochondrial protein initiation by spheroplasts derived from yeast cells in different physiological states and from genetically marked strains. It is low under glucose repression and absent in a strain lacking mitochondrial DNA.