Multiscale Water Dynamics on Protein Surfaces: Protein-Specific Response to Surface Ions.

Proteins function in crowded aqueous environments, interacting with a diverse range of compounds, and among them, dissolved ions. These interactions are water-mediated. In the present study, we combine field-dependent NMR relaxation (NMRD) and theory to probe water dynamics on the surface of proteins in concentrated aqueous solutions of hen egg-white lysozyme (LZM) and bovine serum albumin (BSA). The experiments reveal that the presence of salts (NaCl or NaI) leads to an opposite ion-specific response for the two proteins: an addition of salt to LZM solutions increases water relaxation rates with respect to the salt-free case, while for BSA solutions, a decrease is observed. The magnitude of the change depends on the ion identity. The developed model accounts for the non-Lorentzian shape of the NMRD profiles and reproduces the experimental data over four decades in Larmor frequency (10 kHz to 110 MHz). It is applicable up to high protein concentrations. The model incorporates the observed ion-specific effects via changes in the protein surface roughness, represented by the surface fractal dimension, and the accompanying changes in the surface water residence times. The response is protein-specific, linked to geometrical aspects of the individual protein surfaces, and goes beyond protein-independent Hofmeister-style ordering of ions.

Ion-specificity and surface water dynamics in protein solutions.

Ion-specific effects at the protein surface are investigated here in light of the changes they infer to surface water dynamics, as observed by 1H NMR relaxation (at 20 MHz). Two well-known proteins, hen egg-white lysozyme (LZM) and bovine serum albumin (BSA), show qualitatively opposite trends in the transverse relaxation rate, R2(1H), along a series of different monovalent salt anions in the solution. Presence of salt ions increases R2(1H) in the case of lysozyme and diminishes it in the case of BSA. The effect magnifies for larger and more polarizable ions. The same contrasting effect between the two proteins is observed for protein-solvent proton exchange. This hints at subtle effects ion-binding might have on the accessibility of water surface sites on the protein. We suggest that the combination of the density of surface charge residues and surface roughness, at the atomic scale, dictates the response to the presence of salt ions and is proper to each protein. Further, a dramatic increase in R2(1H) is found to correlate closely with the formation of protein aggregates. The same ordering of salts in their ability to aggregate lysozyme, as seen previously by cloud point measurements, is reproduced here by R2(1H). 1H NMR relaxation data is supplemented by 35Cl and 14N NMR relaxation for selected salt ions to probe the ion-binding itself.

Calorimetric studies of interactions between low molecular weight salts and bovine serum albumin in water at pH values below and above the isoionic point.

Isothermal titration calorimetry was used to determine the temperature and salt concentration dependence of the enthalpy of mixing, Δmix H, of bovine serum albumin (BSA) in aqueous buffer solutions with several low molecular weight salts. Three buffers were used: acetate (pH = 4.0), MOPS (7.2), and borate (9.2). Since the isoionic point of BSA is at pI ≈ 4.7, the net charge of BSA in acetate buffer was positive (≈ +20), while in the other two buffer solutions it was negative (≈ -15 in MOPS and ≈ -25 in borate). The majority of the recorded heat effects were exothermic, while only at pH = 9.2 a weak endothermic effect upon mixing BSA with LiCl, NaCl, and KCl was observed. For all buffer solutions the absolute values of Δmix H of sodium salts followed the order: NaCl < NaBr < NaNO3 < NaI < NaSCN, which is the reverse Hofmeister series for anions. The magnitude of the effects was the largest in acetate buffer and decreased with an increasing pH value of the solution. While the effect of varying the anion of the added salts was strongly pronounced at all pH values, the effect of the cation (LiCl, NaCl, KCl, RbCl and CsCl salts) was weak. The most interesting feature of the results obtained for pH > pI was the fact that Δmix H were considerably more sensitive to the anion (co-ion to the net BSA charge) than to the cation species. This indicated that anions interacted quite strongly with the BSA even at pH values where the net charge of the protein was negative. We showed that Δmix H at high addition of salts correlated well with the enthalpy of hydration of the corresponding salt anion. This finding suggested, consistently with some previous studies, that a part of the exothermic contribution to Δmix H originated from the hydration changes upon the protein-salt interaction. Theoretical analysis, based on the primitive model of highly asymmetric electrolyte solutions solved within the mean spherical approximation, was used to estimate Coulomb effects upon mixing.