Construction and characterization of an Escherichia coli mutant deficient in the metY gene encoding tRNA(f2Met): either tRNA(f1Met) or tRNA(f2Met) is required for cell growth.
The Escherichia coli metY gene, encoding tRNA(f2Met), was split by the kanamycin-resistance-encoding gene. The resulting mutant exhibited the same growth rate as the wild type, indicating that tRNA(f2Met) is not indispensable as is the case with the metZ gene encoding tRNA(f1Met) [Kenri et al., Gene 103 (191) 31-36]. beta-Galactosidase was produced efficiently from the start codon AUG of the intact lacZ gene or a trpA'::lac'Z fusion gene, in the metY mutant. The lac repressor from the lacI gene and the chimeric protein from a hupB'::lac'Z fusion gene, whose start codons are GUG, were also synthesized efficiently in the insertion mutant. These results provide evidence that tRNA(f2Met) is not essential for growth of E. coli and that the start codons, AUG and GUG, are both recognized by tRNA(f1Met), a major N-formyl methionine-specific tRNA, in the tRNA(f2Met)-depleted cells. We were unable to construct mutants deficient in both tRNA(f1Met) and tRNA(f2Met) by P1 phage-mediated transduction with the metY and metZ mutations. Moreover, the ampicillin-resistance marker of the pUC9 plasmid carrying metZ+ was not cured at 42 degrees C in host cells with the polAts and metY-metZ double mutations. These results indicate that either tRNA(f1Met) or tRNA(f2Met) is required for the growth of E. coli.
Kenri
T
T
Department of Molecular Genetics, Kyoto Pharmaceutical University, Japan.
Imamoto
F
F
Kano
Y
Y
D015139:Blotting, Southern; D003062:Codon / Q000235:genetics; D004274:DNA, Recombinant / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D007613:Kanamycin Resistance / Q000235:genetics; D016254:Mutagenesis, Insertional; D009154:Mutation / Q000235:genetics; D010957:Plasmids / Q000235:genetics; D012329:RNA, Bacterial / Q000235:genetics* / Q000378:metabolism; D012358:RNA, Transfer, Met / Q000235:genetics* / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000235:genetics; D001616:beta-Galactosidase / Q000235:genetics
D003062:Codon; D004274:DNA, Recombinant; D012329:RNA, Bacterial; D012358:RNA, Transfer, Met; D011993:Recombinant Fusion Proteins; D001616:beta-Galactosidase