Initiation factors IF1 and IF2 synergistically remove peptidyl-tRNAs with short polypeptides from the P-site of translating Escherichia coli ribosomes

• Published in: Journal of molecular biology Vol. 281; no. 2; pp. 241 - 252
• Main Authors: Karimi, Reza, Pavlov, Michael Yu, Heurgué-Hamard, Valérie, Buckingham, Richard H., Ehrenberg, Måns
• Format: Journal Article
• Language: English
• Published: England Elsevier Ltd 14-08-1998
• Subjects: Carboxylic Ester Hydrolases - metabolism; drop-off; Escherichia coli; Escherichia coli - genetics; Escherichia coli - growth & development; Guanosine Triphosphate - physiology; initiation factors; Oligopeptides - metabolism; Peptide Initiation Factors - metabolism; Peptide Initiation Factors - pharmacology; peptidyl-tRNAs; Prokaryotic Initiation Factor-2; Protein Biosynthesis - physiology; ribosomal P-site; Ribosomes - metabolism; RNA, Transfer, Amino Acyl - metabolism; RNA, Transfer, Met - metabolism; translation; ribosomal P-site; DTE; Pth; PMSF; drop-off; translation; PEP; PK; peptidyl-tRNAs; initiation factors; MK; ERRORS; SUPPRESSION; PROTEIN-SYNTHESIS; DISSOCIATION; PURIFICATION; GENE-EXPRESSION; A-SITE; TRANSFER-RNA HYDROLASE; BINDING
• ISSN: 0022-2836; 1089-8638
• DOI: 10.1006/jmbi.1998.1953

A novel function of initiation factors IF1 and IF2 in
Escherichia coli translation has been identified. It is shown that these factors efficiently catalyse dissociation of peptidyl-tRNAs with polypeptides of different length from the P-site of
E. coli ribosomes, and that the simultaneous presence of both factors is required for induction of drop-off. The factor-induced drop-off occurs with both sense and stop codons in the A-site and competes with peptide elongation or termination. The efficiency with which IF1 and IF2 catalyse drop-off decreases with increasing length of the nascent polypeptide, but is quite significant for hepta-peptidyl-tRNAs, the longest polypeptide chains studied. In the absence of IF1 and IF2 the rate of drop-off varies considerably for different peptidyl-tRNAs, and depends both on the length and sequence of the nascent peptide. Efficient factor-catalysed drop-off requires GTP but not GTP hydrolysis, as shown in experiments without guanine nucleotides, with GDP or with the non-cleavable analogue GMP-PNP.
Simultaneous overexpression of IF1 and IF2
in vivo inhibits cell growth specifically in some peptidyl-tRNA hydrolase deficient mutants, suggesting that initiation factor-catalysed drop-off of peptidyl-tRNA can occur on a significant scale in the bacterial cell. Consequences for the bacterial physiology of this previously unknown function of IF1 and IF2 are discussed.