Two novel methyltransferases acting upon eukaryotic elongation factor 1A in Saccharomyces cerevisiae.
Eukaryotic elongation factor 1A (eEF1A) is an abundant cytosolic protein in Saccharomyces cerevisiae and is well conserved amongst species. This protein undergoes multiple posttranslational modifications, including the N-methylation of four side chain lysine residues. However, the enzyme(s) responsible for catalyzing these modifications have remained elusive. Here we show by intact protein mass spectrometry that deletion of either of two genes coding for putative methyltransferases results in a loss in mass of eEF1A. Deletion of the YHL039W gene, a member of the SET domain subfamily including cytochrome c and ribosomal protein lysine methyltransferases, results in an eEF1A mass loss corresponding to a single methyl group. Deletion in the YIL064W/SEE1 gene, encoding a well conserved seven beta strand methyltransferase sequence, has been shown previously to affect vesicle transport; in this work we show that deletion results in the loss of two methyl group equivalents from eEF1A. We find that deletion of thirty-five other putative and established SET domain and seven beta strand methyltransferases has no effect on the mass of eEF1A. Finally, we show that wild type extracts, but not YIL064W/SEE1 mutant extracts, can catalyze the S-adenosylmethionine-dependent in vitro methylation of hypomethylated eEF1A. We suggest that YHL039W (now designated EFM1 for elongation factor methyltransferase 1) and YIL064W/SEE1 encode distinct eEF1A methyltransferases that respectively monomethylate and dimethylate this protein at lysine residues.
Lipson
Rebecca S
RS
Department of Chemistry and Biochemistry, Molecular Biology Institute, UCLA, Los Angeles, CA 90095-1569, USA.
Webb
Kristofor J
KJ
Clarke
Steven G
SG
D000595:Amino Acid Sequence; D001665:Binding Sites; D017124:Conserved Sequence; D017353:Gene Deletion; D005800:Genes, Fungal; D008239:Lysine / Q000737:chemistry; D008745:Methylation; D008780:Methyltransferases / Q000235:genetics / Q000378:metabolism*; D008969:Molecular Sequence Data; D020648:Peptide Elongation Factor 1 / Q000737:chemistry / Q000378:metabolism*; D011496:Protein Methyltransferases / Q000235:genetics / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012441:Saccharomyces cerevisiae / Q000235:genetics / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins / Q000235:genetics / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid
D020648:Peptide Elongation Factor 1; D029701:Saccharomyces cerevisiae Proteins; D008780:Methyltransferases; D011496:Protein Methyltransferases; C586424:See1 protein, S cerevisiae; D008239:Lysine