Empirical Optimization of Interactions between Proteins and Chemical Denaturants in Molecular Simulations

Abstract

Chemical denaturants are the most commonly used perturbation applied to study protein stability and folding kinetics, as well as the properties of unfolded polypeptides. We build on recent work balancing the interactions of proteins and water, and accurate models for the solution properties of urea and guanidinium chloride, to develop a combined force field which is able to capture the strength of interactions between proteins and denaturants. We use solubility data for a model tetraglycine peptide in each denaturant to tune the protein-denaturant interaction by a novel simulation methodology. We validate the results against data for more complex sequences: single molecule Förster resonance energy transfer data for a 34-residue fragment of the globular protein CspTm, and photoinduced electron transfer quenching data for the disordered peptides C(AGQ)nW in denaturant solution, as well as the chemical denaturation of the mini-protein Trp cage. The combined force field model should aid our understanding of denaturation mechanisms and the interpretation of experiment.

Probing the Action of Chemical Denaturant on an Intrinsically Disordered Protein by Simulation and Experiment

“…Such extensive trajectories, while still posing a challenge for the large systems considered, are the minimum necessary to obtain a representative sampling, given that the experimental reconfiguration times of unfolded and disordered proteins are typically of the order of 0.05–0.1 µs 17, 75–77 . We use force field models for protein, urea and water which we have recently parameterized to reproduce the balance of interactions between the protein, water, and denaturant components of the system 44, 55 . We note that using such a force field is essential, because recent work has shown that most existing force fields result in too collapsed conformations of proteins even in the absence of denaturant 78–79 , with several suggested corrections proposed 55, 80–81 .…”

“…A high local density of water dipoles helps to solvate the charged side-chains, and indeed we observe a very large first peak in the water g(r) around Asp, relative to Asn (Figure S4): This higher water density is a manifestation of the well-known electrostriction effect of ions. Since there are no residues with aromatic side chains in ACTR, yet these usually have the most favorable water-urea transfer free energies in experiment 95 , we have calculated the preferential interaction coefficients of the unfolded Trp cage mini protein using published simulations with the same force field in 3M urea 44 (Figure S5). We find qualitatively that Trp and Tyr have much larger preferential interaction coefficients and therefore more favorable transfer free energy in Trp cage, consistent with experiment.…”

“…Simulations have demonstrated their value in interpreting scattering data on unfolded proteins at large scattering angles, where analytical models may be insufficient 43 . Molecular simulations have already been used in a large number of studies to determine the effect of chemical denaturants on protein stability 5, 44–45 , on the unfolded state 44, 46–47 and on the mechanism by which they denature proteins 4–5, 8, 46, 48–53 . However, the rapid denaturation observed in some of these studies suggested that many of the force fields may not quantitatively capture the effect of denaturants on protein stability.…”

Inferring Properties of Disordered Chains From FRET Transfer Efficiencies

“…A full list of the simulations and proteins used is given in Table S1, with details in Supporting methods. These force fields have been specifically developed with the aim of accurately representing ensembles of unfolded and disordered proteins, and have been validated in several independent studies 36,59,63,66,67 . We therefore believe that these simulations generate more realistic conformational ensembles of unfolded/intrinsically disordered proteins than most other explicit solvent force fields, which typically yield structures that are too collapsed 68 .…”

Perspective: Chain dynamics of unfolded and intrinsically disordered proteins from nanosecond fluorescence correlation spectroscopy combined with single-molecule FRET

“…Triggered primarily by the increasing interest in IDPs, recent developments have changed this situation considerably. 7,11,12,[102][103][104][105][106] Another important development has been the use of experimental restraints, where a simulated conformational ensemble is biased or reweighted based on experimental information, taking into account the uncertainties of the measurements. 85,[107][108][109] These developments have started to enable a more and more realistic and detailed view of unfolded proteins.…”