Regulation of elongation factor G GTPase activity by the ribosomal state. The effects of initiation factors and differentially bound tRNA, aminoacyl-tRNA, and peptidyl-tRNA.

The elongation factor G (EF-G) is responsible for the translocation of the ribosome along the mRNA chain. Under in vitro conditions, EF-G exhibits a very active uncoupled GTPase activity which is dependent on the presence of ribosomes and is modulated by mRNA-dependent binding of tRNA. In the absence of tRNA, uncoupled EF-G GTPase is inhibited by initiation factors IF1 and IF3, but not by initiation factor IF2. In the presence of N-fMet-tRNAfMet and poly(A,U,G) or in the presence of N-acetyl-Phe-tRNAPhe and poly(U), initiation factor IF2 causes an additional decrease of the uncoupled EF-G GTPase activity. This effect, however, is dependent on the presence of IF1 and IF3 and is obviously due to the mRNA- and initiation factor-dependent binding of N-fMet-tRNAfMet and N-acetyl-Phe-tRNAPhe, respectively, to the ribosomal P-site. Non-enzymatic binding of N-fMet-tRNAfMet and N-acetyl-Phe-tRNAPhe, however, causes a stimulation of uncoupled EF-G GTPase activity. The same effects are observed for Met-tRNA, Phe-tRNAPhe and uncharged tRNA. These findings are discussed in the light of the three-site model of the ribosome and the mechanism of translocation.