FEPs <i>V(γ)</i> for <i>γ₃₈₇</i> in <i>k_BT</i> unit.

<p><i>V(γ)</i> computed from APO, ADP-bound and ATP-bound MD simulations of DnaK are colored in red, blue and green respectively.</p

Decipher the Mechanisms of Protein Conformational Changes Induced by Nucleotide Binding through Free-Energy Landscape Analysis: ATP Binding to Hsp70

<div><p>ATP regulates the function of many proteins in the cell by transducing its binding and hydrolysis energies into protein conformational changes by mechanisms which are challenging to identify at the atomic scale. Based on molecular dynamics (MD) simulations, a method is proposed to analyze the structural changes induced by ATP binding to a protein by computing the effective free-energy landscape (FEL) of a subset of its coordinates along its amino-acid sequence. The method is applied to characterize the mechanism by which the binding of ATP to the nucleotide-binding domain (NBD) of Hsp70 propagates a signal to its substrate-binding domain (SBD). Unbiased MD simulations were performed for Hsp70-DnaK chaperone in nucleotide-free, ADP-bound and ATP-bound states. The simulations revealed that the SBD does not interact with the NBD for DnaK in its nucleotide-free and ADP-bound states whereas the docking of the SBD was found in the ATP-bound state. The docked state induced by ATP binding found in MD is an intermediate state between the initial nucleotide-free and final ATP-bound states of Hsp70. The analysis of the FEL projected along the amino-acid sequence permitted to identify a subset of 27 protein internal coordinates corresponding to a network of 91 key residues involved in the conformational change induced by ATP binding. Among the 91 residues, 26 are identified for the first time, whereas the others were shown relevant for the allosteric communication of Hsp70 s in several experiments and bioinformatics analysis. The FEL analysis revealed also the origin of the ATP-induced structural modifications of the SBD recently measured by Electron Paramagnetic Resonance. The pathway between the nucleotide-free and the intermediate state of DnaK was extracted by applying principal component analysis to the subset of internal coordinates describing the transition. The methodology proposed is general and could be applied to analyze allosteric communication in other proteins.</p></div

Statistical analysis of the dissimilarity [<i>1-H(γ_i)</i>] of the FEPs between concatenated MD runs in different nucleotide-binding states.

<p><i>1-H(γ_i)</i>>0.7, 0.3<<i>1-H(γ_i)</i>≤0.7 and <i>1-H(γ_i)</i>≤0.3, respectively. The notation is the following: {number of residues in the subset}/{number of residues compared}. Subset strongly, significantly and weakly influenced correspond to </p

List of the interdomain interactions and their corresponding average distances extracted from the contact map difference ATP/ADP (see text for details).

<p>List of the interdomain interactions and their corresponding average distances extracted from the contact map difference ATP/ADP (see text for details).</p

Theoretical Insights into Sub-Terahertz Acoustic Vibrations of Proteins Measured in Single-Molecule Experiments

Proteins are an important class of nanobioparticles with acoustical modes in the sub-THz frequency range. There is considerable interest to measure and establish the role of these acoustical vibrations for biological function. So far, the technique providing the most detailed information about the acoustical modes of proteins is the very recent Extraordinary Acoustic Raman (EAR) spectroscopy. In this technique, proteins are trapped in nanoholes and excited by two optical lasers of slightly different wavelengths producing an electric field at low frequency (<100 GHz). We demonstrate that the acoustical modes of proteins studied by EAR spectroscopy are both infrared- and Raman-active modes, and we provided interpretation of the spectroscopic fingerprints measured at the single-molecule level. A combination of the present calculations with techniques based on the excitation of a single nanobioparticle by an electric field, such as EAR spectroscopy, should provide a wealth of information on the role of molecular dynamics for biological function

Comparison between MD data and EPR experiments.

<p>Histograms of the distance E430-R547 computed from the MD simulations in the APO state (panels A and C, red) and in the ATP-bound state (panels B and D, green). Panels A and B represent the distributions up to 20 Å, as in the EPR experiments and panels C and D represents the same distributions up to 40 Å. Each subpopulation in the distance distribution was fitted with a Gaussian function shown with black lines. The black arrows in panels A and B represent the experimental results from Ref. <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003379#pcbi.1003379-Schlecht1" target="_blank">[81]</a>. FEP <i>V(γ₅₀₄)</i> computed from different subpopulations relative to the distance E430/R547 distribution function in the APO state (panel E) and in the ATP state (panel F). <i></i> represents the mean value of the Gaussian distributions shown in panel C for the APO-DnaK and D for the ATP-DnaK.</p

Dihedral principal component analysis applied to MD simulations of ATP-bound DnaK.

<p>FES computed for the MD run ATP1. Minima are shown with gray (green for the most probable one) diamonds and the isolines (black lines) are drawn every <i>k_BT</i> unit. The color scale for the free-energy is in <i>k_BT</i> units.</p

List of residues found to be relevant for conformational dynamics of <i>human</i> Hsp70 deduced from NMA of human Hsp70 [49], [92] and from dPCA of <i>E. coli</i> MD trajectories (present work) after sequence alignment of <i>E. coli</i> DnaK on human Hsp70 (Table S1).

<p>Residues common to the two analyses are in bold. Residues of hHsp70 which are not identical in <i>E. coli</i> Hsp70 are in italic font.</p

Analysis of 1-D FEPs of CGDAs <i>γ</i>.

<p>Mapping of the dissimilarity index <i>1-H</i> between the FEPs of APO/ADP (top panel), APO/ATP (middle panel) and ADP/ATP (bottom panel) onto the structure of DnaK (PDB ID: 2KHO) <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003379#pcbi.1003379-Bertelsen1" target="_blank">[70]</a>. Residues belonging to the CGDAs <i>γ</i> which are strongly influenced (<i>1-H</i>>0.7), significantly influenced (0.3≤<i>1-H</i><0.7) and weakly influenced by the nucleotide (<i>1-H</i>≤0.3) are shown with large red, medium yellow and small cyan spheres, respectively. The color code of the protein structure is the same as in <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003379#pcbi-1003379-g001" target="_blank">Fig. 1</a>. These figures were prepared with PyMOL [<a href="http://www.pymol.org" target="_blank">http://www.pymol.org</a>].</p

List of the 27 CGDAs <i>γ</i> revealed by analysis of 1-D FEPs of DnaK in different nucleotide-binding states and their corresponding amino acids.

<p>† and ‡ indicate amino acids which have been depicted to be allosterically relevant from biochemical and bioinformatics methods, respectively. Characters in bold in the table represents strongly and significantly influenced FEPs (<i>H</i><0.7, see <a href="http://www.ploscompbiol.org/article/info:doi/10.1371/journal.pcbi.1003379#pcbi-1003379-t002" target="_blank">Table 2</a> and text for more details). </p