Molecular dynamics simulation of the allosteric regulation of eIF4A protein from the open to closed state, induced by ATP and RNA substrates

Abstract

Background: Eukaryotic initiation factor 4A (eIF4A) plays a key role in the process of protein translation initiation by facilitating the melting of the 5' proximal secondary structure of eukaryotic mRNA for ribosomal subunit attachment. It was experimentally postulated that the closed conformation of the eIF4A protein bound by the ATP and RNA substrates is coupled to RNA duplex unwinding to promote protein translation initiation, rather than an open conformation in the absence of ATP and RNA substrates. However, the allosteric process of eIF4A from the open to closed state induced by the ATP and RNA substrates are not yet fully understood.

Methodology: In the present work, we constructed a series of diplex and ternary models of the eIF4A protein bound by the ATP and RNA substrates to carry out molecular dynamics simulations, free energy calculations and conformation analysis and explore the allosteric properties of eIF4A.

Results: The results showed that the eIF4A protein completes the conformational transition from the open to closed state via two allosteric processes of ATP binding followed by RNA and vice versa. Based on cooperative allosteric network analysis, the ATP binding to the eIF4A protein mainly caused the relative rotation of two domains, while the RNA binding caused the proximity of two domains via the migration of RNA bases in the presence of ATP. The cooperative binding of ATP and RNA for the eIF4A protein plays a key role in the allosteric transition.

Figure 1. Superposition of two complexes.

Superposition of the yeast ATP+RNA+C-eIF4A model (magenta) and the closed human eIF4AIII+ADPNP+RNA complex (gray, PDB-ID 2J0S) in the homology modeling.

Figure 2. RMSD values of the equilibrium and allosteric models.

RMSD values of all backbone atoms with respect to the corresponding starting structures for the CMD simulations of (A) O-eIF4A, (B) ATP+RNA+C-eIF4A, (C) ATP+eIF4A and (D) RNA+eIF4A models, and for the TMD simulations of (E) the (ATP+eIF4A)+RNA→(ATP+RNA+C-eIF4A) transition and (F) the (RNA+eIF4A)+ATP→(ATP+RNA+C-eIF4A) transition.

Figure 3. The structures of the open and closed states.

MD-simulated overall structures for (A) the open eIF4A state and (B) the closed eIF4A state bound by ATP (colored in magenta) and RNA (colored in pink), the linker in which is colored in cyan.

Figure 4. The structures of N- and C- domains of eIF4A.

The structures of (A) the N-domain 5α (yellow) - 8β (orange) - 4α (slate) and (B) the C-domain 2α (yellow) - 7β (orange) - 4α (slate) for the eIF4A protein.

Figure 5. The distances in the open and closed eIF4A states.

The length of whole eIF4A protein (black line), the mass center distance between two domains (magenta line) and the length of the linker (blue line) for (A) the open eIF4A state for the O-eIF4A model and (B) the closed eIF4A state in the ATP+RNA+C-eIF4A model; the α helices and β strands at the N-domain – C-domain interface are labeled and shown in cartoon form with the remaining part of the protein colored in white semi-transparent surface; the ATP and RNA molecules are shown by magenta stick and pink cartoon, respectively, in the closed state.

Figure 6. Domain rotation angel between the O-eIF4A and ATP+eIF4A models.

(A) Dynamic domain identified between the O-eIF4A and ATP+eIF4A models. The residues forming the fixed domain, moving domain and bending residues are depicted in colors of blue, red and green, respectively, with a reference line crossing at the center of rotation. (B) The relative rotation angel of the two domains in the eIF4A protein between the O-eIF4A (slate) and ATP+eIF4A (orange) models.

Figure 7. The structures of the allosteric process.

The average structures are extracted from the trajectories of the O-eIF4A, ATP+eIF4A, I, II, III, IV and ATP+RNA+C-eIF4A models involved in the allosteric process of the ATP binding followed by RNA, i.e., the ATP+eIF4A model for the ATP first binding to the O-eIF4A model; the I-IV models for the RNA second binding to the equilibrium structure of the ATP+eIF4A model; the ATP+RNA+C-eIF4A model for the closed state of the eIF4A protein. The center image shows the time-dependence of the mass center distance between two domains (black), and that of N-domain – C-domain interface (red) during the simulation.

Figure 8. The difference of distances of P-loops.

The difference of the mass center distances of the ATP binding P-loops between the O-eIF4A (slate) and ATP+eIF4A (orange) models.

Figure 9. The distances of RNA binding sites in the eIF4A protein.

The residues (red sticks) at the RNA binding region (α helices and β strands) in (A) the O-eIF4A, (B) ATP+eIF4A and (C) ATP+RNA+C-eIF4A models with the mass center distances of RNA binding sites at the two domains (light blue line).

Figure 10. The fluctuations of residues.

The fluctuations of residues in the O-eIF4A (green), ATP+eIF4A (red) and ATP+RNA+C-eIF4A (blue) models with the labeled α helices and β strands at the ATP/RNA binding region and the N-domain – C-domain interface in the shaded regions.

Figure 11. Correlation maps and the differences of correlations.

Dynamical cross-correlation maps for (A) the allosteric ATP+eIF4A simulation and (B) the differences of motion correlations between the ATP+RNA+C-eIF4A and ATP+eIF4A equilibrium simulations, with specific sub-regions squared in black for the correlations of the ATP binding and RNA binding regions, in magenta for the correlations of the ATP binding region and the N-domain – C-domain interface, in blue for the correlations of the N-domain RNA binding region and the N-domain – C-domain interface, and in green for the correlations of the C-domain RNA binding region and the N-domain – C-domain interface.