Movement of elongation factor G between compact and extended conformations.

Previous structural studies suggested that ribosomal translocation is accompanied by large interdomain rearrangements of elongation factor G (EF-G). Here, we follow the movement of domain IV of EF-G relative to domain II of EF-G using ensemble and single-molecule Förster resonance energy transfer. Our results indicate that ribosome-free EF-G predominantly adopts a compact conformation that can also, albeit infrequently, transition into a more extended conformation in which domain IV moves away from domain II. By contrast, ribosome-bound EF-G predominantly adopts an extended conformation regardless of whether it is interacting with pretranslocation ribosomes or with posttranslocation ribosomes. Our data suggest that ribosome-bound EF-G may also occasionally sample at least one more compact conformation. GTP hydrolysis catalyzed by EF-G does not affect the relative stability of the observed conformations in ribosome-free and ribosome-bound EF-G. Our data support a model suggesting that, upon binding to a pretranslocation ribosome, EF-G moves from a compact to a more extended conformation. This transition is not coupled to but likely precedes both GTP hydrolysis and mRNA/tRNA translocation.