The essential role of the invariant GGQ motif in the function and stability in vivo of bacterial release factors RF1 and RF2.
Release factors RF1 and RF2 are required in bacteria for the cleavage of peptidyl-tRNA. A single sequence motif, GGQ, is conserved in all eubacterial, archaebacterial and eukaryotic release factors and may mimic the CCA end of tRNA, although the position of the motif in the crystal structures of human eRF1 and Escherichia coli RF2 is strikingly different. Mutations have been introduced at each of the three conserved positions. Changing the Gln residue to Ala or Glu allowed the factors to retain about 22% of tetrapeptide release activity in vitro, but these mutants could not complement thermosensitive RF mutants in vivo. None of several mutants with altered Gly residues retained activity in vivo or in vitro. Many GGQ mutants were poorly expressed and are presumably unstable; many were also toxic to the cell. The toxic mutant factors or their degradation products may bind to ribosomes inhibiting the action of the normal factor. These data are consistent with a common role for the GGQ motif in bacterial and eukaryotic release factors, despite strong divergence in primary, secondary and tertiary structure, but are difficult to reconcile with the hypothesis that the amide nitrogen of the Gln plays a vital role in peptidyl-tRNA hydrolysis.
Mora
Liliana
L
UPR9073 du CNRS, Institut de Biologie Physico-Chimique, 13 rue Pierre et Marie Curie, Paris 75005, France.
Heurgué-Hamard
Valérie
V
Champ
Stéphanie
S
Ehrenberg
Måns
M
Kisselev
Lev L
LL
Buckingham
Richard H
RH
D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D029968:Escherichia coli Proteins; D010443:Peptide Chain Termination, Translational; D010454:Peptide Termination Factors / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D015534:Trans-Activators / Q000378:metabolism
D001426:Bacterial Proteins; C489002:ETF1 protein, human; D029968:Escherichia coli Proteins; D010454:Peptide Termination Factors; D015534:Trans-Activators; C041460:peptide chain termination release factor 2; C489005:prfA protein, E coli; C489006:prfB protein, E coli