During the translocation step of the elongation cycle, two
tRNAs together with the mRNA move synchronously and rapidly
on the ribosome. The movement is catalyzed by the binding of
elongation factor G (EF-G) and driven by GTP hydrolysis. Here
we study structural changes of the ribosome related to EF-G
binding and translocation by monitoring the accessibility of
ribosomal RNA (rRNA) for chemical modification by dimethyl sulfate
or cleavage by hydroxyl radicals generated by Fe(II)-EDTA. In
the state of the ribosome that is formed upon binding of EF-G
but before the movement of the tRNAs takes place, residues 1054,
1196, and 1201 in helix 34 in 16S rRNA are strongly protected.
The protections depend on EF-G binding, but do not require GTP
hydrolysis, and are lost upon translocation. Mutants of EF-G,
which are active in ribosome binding and GTP hydrolysis but
impaired in translocation, do not bring about the protections.
According to cryo-electron microscopy (Stark et al., Cell,
2000, 100:301–309), there is no contact of EF-G
with the protected residues of helix 34 in the pretranslocation
state, suggesting that the observed protections are due to an
induced conformational change. Thus, the present results indicate
that EF-G binding to the pretranslocation ribosome induces a
structural change of the head of the 30S subunit that is essential
for subsequent tRNA-mRNA movement in translocation.