Release factor RF3 in E.coli accelerates the dissociation of release factors RF1 and RF2 from the ribosome in a GTP‐dependent manner

Ribosomes complexed with synthetic mRNA and peptidyl‐tRNA, ready for peptide release, were purified by gel filtration and used to study the function of release factor RF3 and guanine nucleotides in the termination of protein synthesis. The peptide‐releasing activity of RF1 and RF2 in limiting concentrations was stimulated by the addition of RF3 and GTP, stimulated, though to a lesser extent, by RF3 and a non‐hydrolysable GTP analogue, and inhibited by RF3 and GDP or RF3 without guanine nucleotide. With short incubation times allowing only a single catalytic cycle of RF1 or RF2, peptide release activity was independent of RF3 and guanine nucleotide. RF3 hydrolysis of GTP to GDP + P i was dependent only on ribosomes and not on RF1 or RF2. RF3 affected neither the rate of association of RF1 and RF2 with the ribosome nor the catalytic rate of peptide release. A model is proposed which explains how RF3 recycles RF1 and RF2 by displacing the factors from the ribosome after the release of peptide.