On the mode of inhibition of eukaryotic protein synthesis by ADP-ribosylation of elongation factor 2

The exchange of flee guanine nucleotides with guanine nucleotides bound to elongation factor 2 (EF-2) and to the EF-2-ribosome complex, and the effect of ADP-ribosylation of the EF-2 thereon, were investigated by nitrocellulo se filter assay. Under the experimental conditions, stoichiometric amounts of guanine nucleotides were bound, in particular, to ternary complexes of E F-2 with biphasic kinetics. The exchange kinetics were similarly biphasic i n all cases. Ribosomes appeared to have variable effects on the exchange ki netics, depending on the type of nucleotide bound. Thus, in their presence, the rate and magnitude of the fast exchange of nucleotides revealed increa sing values in the order GTP (GXP) > GTP gamma S > GDP, ADP-ribosylation ha d no inhibitory effect on the binding of guanine nucleotides to EF-2 or to the EF-2-ribosome complex but reduced significantly the fast exchange of GT P (GXP) and GTP gamma S bound to the EF-2-ribosome complex. The effect of A DP-ribosylation on the fast exchange of GDP in binary and ternary complexes mas less pronounced. The mechanism of inhibition of protein synthesis by A DP-ribosylation of EF-2 is discussed in view of these data.