Simulating movement of tRNA through the ribosome during hybrid-state formation

Abstract

Biomolecular simulations provide a means for exploring the relationship between flexibility, energetics, structure, and function. With the availability of atomic models from X-ray crystallography and cryoelectron microscopy (cryo-EM), and rapid increases in computing capacity, it is now possible to apply molecular dynamics (MD) simulations to large biomolecular machines, and systematically partition the factors that contribute to function. A large biomolecular complex for which atomic models are available is the ribosome. In the cell, the ribosome reads messenger RNA (mRNA) in order to synthesize proteins. During this essential process, the ribosome undergoes a wide range of conformational rearrangements. One of the most poorly understood transitions is translocation: the process by which transfer RNA (tRNA) molecules move between binding sites inside of the ribosome. The first step of translocation is the adoption of a "hybrid" configuration by the tRNAs, which is accompanied by large-scale rotations in the ribosomal subunits. To illuminate the relationship between these rearrangements, we apply MD simulations using a multi-basin structure-based (SMOG) model, together with targeted molecular dynamics protocols. From 120 simulated transitions, we demonstrate the viability of a particular route during P/E hybrid-state formation, where there is asynchronous movement along rotation and tRNA coordinates. These simulations not only suggest an ordering of events, but they highlight atomic interactions that may influence the kinetics of hybrid-state formation. From these simulations, we also identify steric features (H74 and surrounding residues) encountered during the hybrid transition, and observe that flexibility of the single-stranded 3'-CCA tail is essential for it to reach the endpoint. Together, these simulations provide a set of structural and energetic signatures that suggest strategies for modulating the physical-chemical properties of protein synthesis by the ribosome.

Molecular mechanics of 30S subunit head rotation

“…The latter step is accompanied by the intrasubunit rotation of the small-subunit head domain, which unlocks the steric barrier between the P and E sites on the 30S subunit (10-13) and transports the P-site tRNA into the 30S E site (14-16). In addition to a growing structural database of trapped translocation complexes from X-ray and cryo-EM studies (Table S1), computational approaches are being used to analyze ribosome dynamics and to describe the energy landscape of the translocation process (17)(18)(19)(20)(21)(22).Structures of trapped EF-G-containing translocation intermediates show that, during 30S subunit head rotation, the translocating P-site tRNA precisely maintains contact with the head domain but moves relative to the 30S body domain, forming a chimeric hybrid (pe/E) state (14, 15). (Lowercase letters indicate that the tRNA is bound in a chimeric hybrid state, whereas uppercase letters indicate binding to the canonical A, P, or E sites.…”

“…The latter step is accompanied by the intrasubunit rotation of the small-subunit head domain, which unlocks the steric barrier between the P and E sites on the 30S subunit (10-13) and transports the P-site tRNA into the 30S E site (14-16). In addition to a growing structural database of trapped translocation complexes from X-ray and cryo-EM studies (Table S1), computational approaches are being used to analyze ribosome dynamics and to describe the energy landscape of the translocation process (17)(18)(19)(20)(21)(22).…”

Perspective: Reaches of chemical physics in biology

“…In this issue, Whitford and Sanbonmatsu use targeted molecular dynamics to reveal different pathways by which tRNAse interacts with the ribosome during translocation. 88 Such results can provide insight for future experimental control of ribosome dynamics, allowing protein manufacture to be experimentally controlled via dynamics, as opposed to just mRNA sequence.…”

The translation elongation cycle—capturing multiple states by cryo-electron microscopy

“…Molecular dynamics simulations of the full ribosome started with the groundbreaking work of Sanbonmatsu et al, who studied the process of aminoacyl-tRNA accommodation in the ribosomal A site [97]. Larger-scaled computations covering increasingly larger time frames have followed since [50,[98][99][100]. For instance, Whitford et al [50] conducted explicit solvent simulations at the atomic level (2.1 million atoms) for 1.3 ms in exploring mRNA-tRNA translocation.…”