FUNCTION OF POLYPEPTIDE-CHAIN RELEASE FACTOR RF-3 IN ESCHERICHIA-COLI- RF-3 ACTION IN TERMINATION IS PREDOMINANTLY AT UGA-CONTAINING STOP SIGNALS

Two protein release factors (RFs) showing codon specificity, RF-1 (UAG , UAA) and RF-2 (UAA, UGA), are required for polypeptide chain termina tion in Escherichia coli. We recently reported the localization and ch aracterization of the gene encoding RF-3 (prfC), a third protein compo nent previously described as stimulating termination without codon spe cificity, RF-3 is a GTP-binding protein that displays much sequence si milarity to elongation factor EF-G. In a termination assay in vitro, R F-3 lowers the K-m for terminator trinucleotides and is thought to act in termination signal recognition. The gene prfC was identified by tr ansposon insertion mutagenesis leading to enhanced nonsense suppressio n of UGA. We report here that (i) RF-3 inactivation significantly enha nces the suppression of termination in vivo only at UGA-dependent stop signals; (ii) the codon dependent contribution to the stimulation of fMet release in vitro by RF-3 is significantly greater with UGA termin ation triplet than UAG termination triplet; (iii) RF-3 increases drama tically the affinity of RF-2 to the UGA termination complex in vitro b ut not that of RF-1 to the UAG termination complex; (iv) RF-3 inactiva tion leads to a positive feedback on the autoregulation of RF-2 synthe sis in vivo, dependent on the competition between frameshifting and te rmination. These findings are discussed in terms of the mechanism of i nvolvement of RF-3 in translation termination.