Protein methylation reactions can play important roles in cell physiology. After labeling intact Saccharomyces cerevisiae cells with S-adenosyl-l-[methyl-(3)H]methionine, we identified a major methylated 49-kDa polypeptide containing [(3)H]methyl groups in two distinct types of linkages. Peptide sequence analysis of the purified methylated protein revealed that it is eukaryotic elongation factor 1A (eEF1A, formerly EF-1alpha), the protein that forms a complex with GTP and aminoacyl-tRNAs for binding to the ribosomal A site during protein translation. Previous studies have shown that eEF1A is methylated on several internal lysine residues to give mono-, di-, and tri-N-epsilon-methyl-lysine derivatives. We confirm this finding but also detect methylation that is released as volatile methyl groups after base hydrolysis, characteristic of ester linkages. In cycloheximide-treated cells, methyl esterified eEF1A was detected largely in the ribosome and polysome fractions; little or no methylated protein was found in the soluble fraction. Because the base-labile, volatile [methyl-(3)H]radioactivity of eEF1A could be released by trypsin treatment but not by carboxypeptidase Y or chymotrypsin treatment, we suggest that the methyl ester is present on the alpha-carboxyl group of its C-terminal lysine residue. From the results of pulse-chase experiments using radiolabeled intact yeast cells, we find that the N-methylated lysine residues of eEF1A are stable over 4 h, whereas the eEF1A carboxyl methyl ester has a half-life of less than 10 min. The rapid turnover of the methyl ester suggests that the methylation/demethylation of eEF1A at the C-terminal carboxyl group may represent a novel mode of regulation of the activity of this protein in yeast.
|Evidence ID||Analyze ID||Interactor||Interactor Systematic Name||Interactor||Interactor Systematic Name||Type||Assay||Annotation||Action||Modification||Phenotype||Source||Reference||Note|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Gene Ontology Term||Gene Ontology Term ID||Qualifier||Aspect||Method||Evidence||Source||Assigned On||Annotation Extension||Reference|
|Evidence ID||Analyze ID||Gene||Gene Systematic Name||Phenotype||Experiment Type||Experiment Type Category||Mutant Information||Strain Background||Chemical||Details||Reference|
|Evidence ID||Analyze ID||Regulator||Regulator Systematic Name||Target||Target Systematic Name||Experiment||Assay||Construct||Conditions||Strain Background||Reference|