H. Aoki et al., THE GENE ENCODING THE ELONGATION-FACTOR-P PROTEIN IS ESSENTIAL FOR VIABILITY AND IS REQUIRED FOR PROTEIN-SYNTHESIS, The Journal of biological chemistry, 272(51), 1997, pp. 32254-32259
Elongation factor P (EFP) is a protein that stimulates the peptidyltra
nsferase activity of fully assembled 70 S prokaryotic ribosomes and en
hances the synthesis of certain dipeptides initiated by N-formylmethio
nine. This reaction appears conserved throughout species and is promot
ed in eukaryotic cells by a homologous protein, eIF5A. Here we ask whe
ther the Escherichia coil gene encoding EFP is essential for cell viab
ility, A kanamycin resistance (Kan(R)) gene was inserted near the N-te
rminal end of the efp gene and was cloned into a plasmid, pMAK705, tha
t has a temperature-sensitive origin of replication. After transformat
ion into a recA(+) E. coli strain, temperature-sensitive mutants were
isolated, and their chromosomal DNA was sequenced. Mutants containing
the efp-Kan(R) gene in the chromosome grew at 33 degrees C only in the
presence of the wild-type copy of the efp gene in the pMAK705 plasmid
and were unable to grow at 44 degrees C. Incorporation of various iso
topes in vivo suggests that translation is impaired in the efp mutant
at 44 degrees C. At 44 degrees C, mutant cells are severely defective
in peptide-bond formation. We conclude that the efp gene is essential
for cell viability and is required for protein synthesis.