Effect of vanadate on EF-G function on the ribosome. (A) Inhibition by vanadate of post-termination complex disassembly. Post-termination complex was rapidly mixed with EF-G/RRF in the absence (1) or presence (2) of vanadate (2 mM), and complex disassembly was monitored by light scattering (LS) (Materials and Methods). (B) Concentration dependence of the inhibition by vanadate. Post-termination complex disassembly was measured as in A in the presence of increasing concentrations of vanadate. Plotted is the half-life (τ1/2) of the reaction measured in the absence relative to that measured in the presence of vanadate; τ1/2 was ∼5 sec in the absence and ∼150 sec in the presence of vanadate. (C) Comparison of post-termination complex disassembly and translocation. Rates of post-termination complex disassembly (black bars) were measured as in A. The rate of single-round translocation (∼15 sec−1, open bars) was measured by fluorescence stopped-flow, multiple-round translocation (hatched bars) by the puromycin assay (Materials and Methods); plotted is the relative initial rate of translocation measured in the absence (2 sec−1, set to 1.0) and presence (0.17 sec−1) of vanadate. The concentration of vanadate was 2 mM.
