title: A novel post-translational modification of yeast elongation factor 1A. Methylesterification at the C terminus.
abstract: 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.
authors.affiliation: Department of Chemistry and Biochemistry and the Molecular Biology Institute, UCLA, Los Angeles, California 90095-1569, USA.
authors.forename: P
authors.initials: P
authors.lastname: Zobel-Thropp
authors.forename: M C
authors.initials: MC
authors.lastname: Yang
authors.forename: L
authors.initials: L
authors.lastname: Machado
authors.forename: S
authors.initials: S
authors.lastname: Clarke
mesh_terms: D002384:Catalysis; D002852:Chromatography, Ion Exchange; D003513:Cycloheximide / Q000494:pharmacology; D006868:Hydrolysis; D008745:Methylation; D008970:Molecular Weight; D020648:Peptide Elongation Factor 1 / Q000096:biosynthesis* / Q000737:chemistry; D011499:Protein Processing, Post-Translational; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012270:Ribosomes / Q000378:metabolism; D012441:Saccharomyces cerevisiae
chemical_list: D020648:Peptide Elongation Factor 1; D011500:Protein Synthesis Inhibitors; D003513:Cycloheximide