Molecular Dynamics Simulations of the Ribosome - Biophysical Approaches to Translational Control of Gene Expression - page 63

structure-based simulations were similar. Both simulations showed agreement with
X-ray B-factors (Korostelev et al. 2006 ), with the exception of the 50S stalks, which
have been shown to undergo large amplitude fluctuations by single molecule FRET
(Munro et al. 2010a, b ) .

These fl uctuations cannot be captured by X-ray

crystallography.

3.4.4 The L1 Stalk

Finally, we used the structure-based method to investigate L1 stalk fluctuations. We
found that enormous fluctuations both in displacement and twist, consistent with the
large fluctuations observed by single molecule FRET.

3.4.5 Moving tRNAs Through the Ribosome

In the case of accommodation, our structure-based simulations display a wide vari-
ety of accommodation pathways. The shape of the pathway distribution is deter-
mined both by the intrinsic properties of the tRNA and by fluctuations of the
ribosome. Furthermore, both the structure-based and explicit solvent simulations
show large ensembles of configurations representing the A/A and A/T positions,
consistent with single molecule FRET experiments. Our study reveals that large
amplitude reversible excursions of the aminoacyl-tRNA combined with stochastic
gating of helix 89 allow for accommodation. For accommodation to occur, the
entropic barrier introduced by the 3¢-CCA end must be overcome by (1) the enthal-
pic minima of the peptidyl transferase center, which includes a base pair between
the 3¢-CCA end and a ribosomal nucleotide, (2) the enthalpic minima of the anti-
codon arm, which releases energy stored during ternary complex binding, and (3) an
entropic “kick” produced by the presence of EF-Tu, which prevents the tRNA from
escaping the ribosome. It should be emphasized that while extremely flexible com-
ponents introduce entropic barriers, they also soften the system relative to a molecu-
lar machine consisting of more rigid components. This softening may act to fine-tune
the free energy landscape. Overall, simulations have established several principles
important for ribosome dynamics: multiple pathways, stochastic gating, large
amplitude reversible excursions, and entropic barriers.
We suspect that similar principles operate during translocation. However, in this
case, the energy landscape seen by the tRNA is itself dynamic: pivoting of the 30S
body with respect to the 50S, swiveling of the 30S head, and the opening and clos-
ing of various gates will significantly alter the barriers in a dynamic fashion. Recent
single molecule FRET experiments suggest that each component of the ribosome
fluctuates independently and at different time scales. It is only during the rare
moments that these fluctuations occur simultaneously in the proper directions that
translocation may occur (Munro et al. 2010a, b ). Such synchronization is enhanced
by the presence of EF-G.