Functional interactions between yeast translation eukaryotic elongation factor (eEF) 1A and eEF3.
The translation elongation machinery in fungi differs from other eukaryotes in its dependence upon eukaryotic elongation factor 3 (eEF3). eEF3 is essential in vivo and required for each cycle of the translation elongation process in vitro. Models predict eEF3 affects the delivery of cognate aminoacyl-tRNA, a function performed by eEF1A, by removing deacylated tRNA from the ribosomal Exit site. To dissect eEF3 function and its link to the A-site activities of eEF1A, we have identified a temperature-sensitive allele of the YEF3 gene. The F650S substitution, located between the two ATP binding cassettes, reduces both ribosome-dependent and intrinsic ATPase activities. In vivo this mutation increases sensitivity to aminoglycosidic drugs, causes a 50% reduction of total protein synthesis at permissive temperatures, slows run-off of polyribosomes, and reduces binding to eEF1A. Reciprocally, excess eEF3 confers synthetic slow growth, increased drug sensitivity, and reduced translation in an allele specific fashion with an E122K mutation in the GTP binding domain of eEF1A. In addition, this mutant form of eEF1A shows reduced binding of eEF3. Thus, optimal in vivo interactions between eEF3 and eEF1A are critical for protein synthesis.
Anand
Monika
M
Department of Molecular Genetics, Microbiology & Immunology, University of Medicine and Dentistry of New Jersey Robert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
Chakraburtty
Kalpana
K
Marton
Matthew J
MJ
Hinnebusch
Alan G
AG
Kinzy
Terri Goss
TG
D000255:Adenosine Triphosphate / Q000378:metabolism; D000483:Alleles; D000617:Aminoglycosides; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001665:Binding Sites; D003001:Cloning, Molecular; D005656:Fungal Proteins / Q000378:metabolism*; D005982:Glutathione Transferase / Q000378:metabolism; D049452:Green Fluorescent Proteins; D006868:Hydrolysis; D008164:Luminescent Proteins / Q000378:metabolism; D009154:Mutation; D020648:Peptide Elongation Factor 1 / Q000378:metabolism*; D010445:Peptide Elongation Factors / Q000378:metabolism*; D010641:Phenotype; D011485:Protein Binding; D014176:Protein Biosynthesis; D017434:Protein Structure, Tertiary; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D012441:Saccharomyces cerevisiae / Q000378:metabolism*; D029701:Saccharomyces cerevisiae Proteins; D013696:Temperature; D013997:Time Factors
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D005656:Fungal Proteins; D008164:Luminescent Proteins; D020648:Peptide Elongation Factor 1; D010445:Peptide Elongation Factors; D011500:Protein Synthesis Inhibitors; D029701:Saccharomyces cerevisiae Proteins; C111584:YEF3 protein, S cerevisiae; D049452:Green Fluorescent Proteins; D000255:Adenosine Triphosphate; D005982:Glutathione Transferase