Evidence for microscopic, long-range hydration forces for a hydrophobic amino acid (hydration structureyprotein folding)

We have combined neutron solution scatter- ing experiments with molecular dynamics simulation to iso- late an excess experimental signal that is caused solely by N-acetyl-leucine-amide (NALA) correlations in aqueous so- lution. This excess signal contains information about how NALA molecule centers are correlated in water, and we show how these solute-solute correlations might be determined at dilute concentrations in the small angle region. We have tested qualitatively different pair distribution functions for NALA molecule centers—gas, cluster, and aqueous forms of gc(r)— and have found that the excess experimental signal is ade- quate enough to rule out gas and cluster pair distribution functions. The aqueous form of gc(r) that exhibits a solvent- separated minimum, and possibly longer-ranged correlations as well, is not only physically sound but reproduces the experimental data reasonably well. This work demonstrates that important information in the small angle region can be mined to resolve solute-solute correlations, their lengthscales, and thermodynamic consequences even at dilute concentra- tions. The hydration forces that operate on the microscopic scale of individual amino acid side chains, implied by the small angle scattering data, could have significant effects on the early stages of protein folding, on ligand binding, and on other intermolecular interactions. Energy landscape models have defined a ''new'' view of protein folding for explaining the kinetics and thermodynamics of protein folding (1-3). The free energy surface is postulated to be funnel-like in shape; that is, the energy decreases faster than the diminishment in the number of states, but with a folded structure minimum that is unique and well separated in energy from the nearest non-native state. Both long and short-ranged forces are important because both imply average funnel-like behavior whereas the latter ensure the uniqueness of the native structure minimum. These theoretical conclusions are partly based on highly idealized lattice models of proteins, which have no atomic detail of amino acid side chains and use very nonspecific descriptions of residue-residue interactions, and where individual beads are considered to be several amino acids that have been ''renormalized'' (1-3). Although the concept of funnel-like energy landscapes is an appealing one, no definitive connection has been made between the landscape model and the genuine physical forces such as hydrogen- bonding or hydration, etc., that may actually give rise to a funneled energy surface. What is the molecular origin of these free energy biases, and how do we determine them? Our intuition is that amino acid interactions mediated by aqueous solvent are a dominant feature of funneled landscapes in protein folding. We have been especially interested in the idea that solute molecules may influence the structure of water out to a distance of several hydration layers from the surface and that these alterations in water structure may in turn give rise to microscopic long- ranged favorable or unfavorable hydration forces between hydrophobic and hydrophilic solutes, respectively (4-7). We define microscopic long-range hydration forces to mean a significant free energy stabilization of amino acid groups in water beyond the point at which they are in van der Waals contact. This contrasts with simpler hydration models based on minimizing hydrophobic solvent-accessible surface area. The estimation of the range and magnitude of microscopic hydration forces acting between amino acids will require the development of an approach that is sensitive to both water structure and any thermodynamic forces present because of hydration. Analysis of neutron solution scattering experiments of amino acids in water by using molecular dynamics simula- tions probed changes in water structure arising from a shift of the main water diffraction peak (5-7). In this paper, we have combined information from experiments and simulation to isolate N-acetyl-leucine-amide (NALA) correlations in water found in the small angle region. After subtracting the simu- lated terms from the total measured scattering, we isolated scattering caused by NALA correlations and determined a model, gc(r), that best reproduces this excess signal. Once the solute-solute pair correlation function in aqueous solution was determined, it could be related to hydration forces through