Urea–Water Solvation Forces on Prion Structures

Solvation forces are crucial determinants in the equilibrium between the folded and unfolded state of proteins. Particularly interesting are the solvent forces of denaturing solvent mixtures on folded and misfolded states of proteins involved in neurodegeneration. The C-terminal globular domain of the ovine prion protein (1UW3) and its analogue H2H3 in the α-rich and β-rich conformation were used as model structures to study the solvation forces in 4 M aqueous urea using molecular dynamics. The model structures display very different secondary structures and solvent exposures. Most protein atoms favor interactions with urea over interactions with water. The force difference between protein–urea and protein–water interactions correlates with hydrophobicity; i.e., urea interacts preferentially with hydrophobic atoms, in agreement with results from solvent transfer experiments. Solvent Shannon entropy maps illustrate the mobility gradient of the urea–water mixture from the first solvation shell to the bulk. Single urea molecules replace water in the first solvation shell preferably at locations of relatively high solvent entropy

Characterisation of HOIP RBR E3 ligase conformational dynamics using integrative modelling

Multidomain proteins composed of individual domains connected by flexible linkers pose a challenge for structural studies due to their intrinsic conformational dynamics. Integrated modelling approaches provide a means to characterise protein flexibility by combining experimental measurements with molecular simulations. In this study, we characterise the conformational dynamics of the catalytic RBR domain of the E3 ubiquitin ligase HOIP, which regulates immune and inflammatory signalling pathways. Specifically, we combine small angle X-ray scattering experiments and molecular dynamics simulations to generate weighted conformational ensembles of the HOIP RBR domain using two different approaches based on maximum parsimony and maximum entropy principles. Both methods provide optimised ensembles that are instrumental in rationalising observed differences between SAXS-based solution studies and available crystal structures and highlight the importance of interdomain linker flexibility

The C terminal domain of troponin C in complex with EGCg and EMD 57033.

<p>Left: cCTnC in complex with EGCg. Calcium ions are shown in mauve. The residues Met120, Leu121, Leu136, Met157 and Val160 are in orange, Gly159 in yellow. The solution NMR binding pose for EGCg (model M) is in violet, <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070556#pone.0070556-Robertson1" target="_blank">[13]</a> while the docked poses corresponding to models M, M and M are respectively in grey, red and cyan. Right: cCTnC in complex with EMD 57033 <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0070556#pone.0070556-Li1" target="_blank">[25]</a> (model M).</p

Average number of the inter-protein hydrogen bonds and salt bridges between cCTnC2Ca and cTnI(34-71) (wild type and with the Gly159Asp mutation) in the presence and absence of EGCg.

<p>The percentage of salt bridges with respect to the total number of hydrogen bonds is shown in brackets. Replica simulations are shown on consecutive rows.</p

Enthalpic and entropic contributions and binding free energies between the complexes in square brackets evaluated within the MM/PBSA and normal mode analysis scheme.

<p>The top part of the table estimates the binding of EGCg or EMD 57033 to cCTnC; the middle part estimates the binding of EGCg when the anchoring part of cTnI is present; the bottom part of the table evaluates the inter-protein binding between the anchoring part of cTnI with cCTnC in the investigated models. Replica simulations are shown on consecutive rows.</p

The structures of EGCg and EMD 57033.

<p>Top: the green tea polyphenol EGCg with labels for carbon bonded hydrogen atoms (left), and rings and dihedral angles (right). Bottom: the calcium sensitiser EMD 57033.</p

Distribution of charged amino acid in cCTnC2CaTnI(34-71).

<p>Molecular dynamics snapshot of cCTnC2CaTnI(34–71) interacting with EGCg coloured according to residue type: non-polar (white), polar (green), positively charged (blue) and negatively charged (red).</p

EGCg in complex with cCTnC when the anchoring region of cTnI is present.

<p>Molecular dynamics snapshots of cCTnC2CaTnI(34–71), wild type (left) and with the Gly159Asp mutation (right), interacting with EGCg. cCTnC is shown in green and cTnI(34–71) in red. The cCTnC residue at position 159 is in blue. The cCTnC residues which formed on average more than 0.4 hydrogen bonds with EGCg are in orange.</p

Specialized Dynamical Properties of Promiscuous Residues Revealed by Simulated Conformational Ensembles

The ability to interact with different partners is one of the most important features in proteins. Proteins that bind a large number of partners (hubs) have been often associated with intrinsic disorder. However, many examples exist of hubs with an ordered structure, and evidence of a general mechanism promoting promiscuity in ordered proteins is still elusive. An intriguing hypothesis is that promiscuous binding sites have specific dynamical properties, distinct from the rest of the interface and pre-existing in the protein isolated state. Here, we present the first comprehensive study of the intrinsic dynamics of promiscuous residues in a large protein data set. Different computational methods, from coarse-grained elastic models to geometry-based sampling methods and to full-atom Molecular Dynamics simulations, were used to generate conformational ensembles for the isolated proteins. The flexibility and dynamic correlations of interface residues with a different degree of binding promiscuity were calculated and compared considering side chain and backbone motions, the latter both on a local and on a global scale. The study revealed that (a) promiscuous residues tend to be more flexible than nonpromiscuous ones, (b) this additional flexibility has a higher degree of organization, and (c) evolutionary conservation and binding promiscuity have opposite effects on intrinsic dynamics. Findings on simulated ensembles were also validated on ensembles of experimental structures extracted from the Protein Data Bank (PDB). Additionally, the low occurrence of single nucleotide polymorphisms observed for promiscuous residues indicated a tendency to preserve binding diversity at these positions. A case study on two ubiquitin-like proteins exemplifies how binding promiscuity in evolutionary related proteins can be modulated by the fine-tuning of the interface dynamics. The interplay between promiscuity and flexibility highlighted here can inspire new directions in protein–protein interaction prediction and design methods

Superimposition of the Phe-bound prephenate dehydratase (PDT) and the tetrameric model of the hPAH.

<p>The Phe-bound PDT from <i>Chlorobium tepidum TLS</i> (PDB code 2QMX) is drawn in cyan. The RD of a subunit of hPAH is drawn in green, whereas the catalytic domain of the adjacent subunit is drawn in violet. The bound Phe ligand is shown as a yellow stick.</p