Cooperative water filling of a nonpolar protein cavity observed by high-pressure crystallography and simulation

247

243

To better understand the role of surface chemical heterogeneity in natural nanoscale hydration, we study via molecular dynamics simulation the structure and thermodynamics of water confined between two protein-like surfaces. Each surface is constructed to have interactions with water corresponding to those of the putative hydrophobic surface of a melittin dimer, but is flattened rather than having its native ''cupped'' configuration. Furthermore, peripheral charged groups are removed. Thus, the role of a rough surface topography is removed, and results can be productively compared with those previously observed for idealized, atomically smooth hydrophilic and hydrophobic flat surfaces. The results indicate that the protein surface is less hydrophobic than the idealized counterpart. The density and compressibility of water adjacent to a melittin dimer is intermediate between that observed adjacent to idealized hydrophobic or hydrophilic surfaces. We find that solvent evacuation of the hydrophobic gap (cavitation) between dimers is observed when the gap has closed to sterically permit a single water layer. This cavitation occurs at smaller pressures and separations than in the case of idealized hydrophobic flat surfaces. The vapor phase between the melittin dimers occupies a much smaller lateral region than in the case of the idealized surfaces; cavitation is localized in a narrow central region between the dimers, where an apolar amino acid is located. When that amino acid is replaced by a polar residue, cavitation is no longer observed.confinement ͉ water ͉ biopolymer ͉ compressibility ͉ cavitation I t has long been accepted that the hydrophobic effect plays a key role in the stability of compact native protein structures (1-3). The protein contains hydrophobic regions which associate, at least in part, due to the favorable solvent-mediated free energy of aggregation of nonpolar moieties in an aqueous environment (2, 4, 5). Although experimental studies (6) suggest that this rationalization is valid, and theoretical work using model systems and realistic protein structures (7) confirms such observations, much is still unknown about the role and behavior of water near proteins and how the aqueous solvent contributes to protein structural stability at the molecular level. The main focus of the present work is to contribute to the understanding of water near, and between, nominally hydrophobic, but realistic, protein surfaces.When one turns to the molecular details of the mechanism of nonpolar aggregation in water, the picture is still not completely clear. The two limiting scenarios for events such as protein folding and directed self-assembly are summarized well in ref. 8, in the context of protein folding. The basic feature distinguishing these scenarios is the relationship between solute dehydration and solute spatial approach. In the traditional view, water is gradually reduced within and between the associating regions in a manner that is concerted with their spatial approach. In an alternative cavitation sce...

“…4a indicates approximately the region of cavitation between the dimers. The size of this region is comparable to that of nonpolar protein cavities found in experiments (36,37). Fig.…”

Section: Resultssupporting

confidence: 82%

Hydrophobicity of protein surfaces: Separating geometry from chemistry

Giovambattista

López

Rossky

et al. 2008

247

243

“…Agreement between free energy calculations and x-ray crystallography was also found for a 160 Å 3 nonpolar cavity engineered into T4 lysozyme, which was found to be empty under ambient conditions but cooperatively filled by several water molecules at kbar pressures (23).…”

mentioning

confidence: 59%

A dry ligand-binding cavity in a solvated protein

Qvist

Davidovic

Hamelberg

et al. 2008

129

Ligands usually bind to proteins by displacing water from the binding site. The affinity and kinetics of binding therefore depend on the hydration characteristics of the site. Here, we show that the extreme case of a completely dehydrated free binding site is realized for the large nonpolar binding cavity in bovine ␤-lactoglobulin. Because spatially delocalized water molecules may escape detection by x-ray diffraction, we use water 17 O and 2 H magnetic relaxation dispersion (MRD), 13 C NMR spectroscopy, molecular dynamics simulations, and free energy calculations to establish the absence of water from the binding cavity. Whereas carbon nanotubes of the same diameter are filled by a hydrogenbonded water chain, the MRD data show that the binding pore in the apo protein is either empty or contains water molecules with subnanosecond residence times. However, the latter possibility is ruled out by the computed hydration free energies, so we conclude that the 315 Å 3 binding pore is completely empty. The apo protein is thus poised for efficient binding of fatty acids and other nonpolar ligands. The qualitatively different hydration of the ␤-lactoglobulin pore and carbon nanotubes is caused by subtle differences in water-wall interactions and water entropy.␤-lactoglobulin ͉ free energy simulation ͉ hydrophobic hydration ͉ magnetic relaxation dispersion G lobular proteins are stabilized by dense atomic packing and by water exclusion from the nonpolar core region. However, the packing density is not uniform and a typical protein contains approximately four cavities per 100 residues of sufficient size to accommodate at least one water molecule (1). Small nonpolar cavities are usually empty and can be regarded as packing defects, whereas small polar cavities tend to be occupied by structural water molecules that stabilize the folded protein by H-bonding to otherwise unsatisfied peptide partners (2). Large cavities are frequently linked to protein functions, such as ligand binding and transport, membrane translocation, and enzyme catalysis. Some of these large cavities are lined exclusively or predominantly by nonpolar sidechains. The question whether such large nonpolar cavities are empty or contain water molecules is of fundamental biological importance (3-8).The principal tool of structural biology, x-ray crystallography, cannot easily resolve this issue. There are three potential problems. (i) Because water molecules interact only weakly with the nonpolar cavity walls, they tend to be positionally disordered. As a result of such delocalization, the electron density of the water molecules may fall below the detection limit. (ii) At a typical resolution limit of Ϸ2 Å, a contaminating nonpolar ligand may be misinterpreted as a water cluster (9, 10). (iii) For structures determined at cryogenic temperatures, the hydration status of the cavity may be altered by re-equilibration during the flash cooling process (11).In favorable cases, water molecules in nonpolar protein cavities can be inferred from intermolecular nuclear O...

“…Water ''evaporating'' from nonpolar confinement has been observed in a number of simulation studies and has been unequivocally demonstrated in at least 1 experiment probing the hydration of a protein cavity (49). Simulation examples of such confinement-induced drying (50) range from nanotubes (51) to plates (28,45,(52)(53)(54)(55)(56)(57)(58)(59) and the interfaces between proteins (30,60) to collapsing polymers (25)(26)(27).…”

Section: Discussionmentioning

confidence: 98%

Static and dynamic correlations in water at hydrophobic interfaces

Mittal

Hummer

2008

Self Cite

137

144

We study the static and dynamic properties of the water-density fluctuations in the interface of large nonpolar solutes. With the help of extensive molecular dynamics simulations of TIP4P water near smooth spherical solutes, we show that for large solutes, the interfacial density profile is broadened by capillary waves. For purely repulsive solutes, the squared width of the interface increases linearly with the logarithm of the solute size, as predicted by capillary-wave theory. The apparent interfacial tension extracted from the slope agrees with that of a free liquid-vapor interface. The characteristic length of local density fluctuations is Ϸ0.5 nm, measured along the arc, again consistent with that of a free liquid-vapor interface. Probed locally, the interfacial density fluctuations exhibit large variances that exceed those expected for an ideal gas. Qualitatively consistent with theories of the free liquid-vapor interface, we find that the water interface near large and strongly nonpolar solutes is flickering, broadened by capillarywave fluctuations. These fluctuations result in transitions between locally wet and dry regions that are slow on a molecular time scale.capillary waves ͉ drying transition ͉ hydrophobic effect ͉ surface tension T he hydration structure and thermodynamics of simple nonpolar solutes in water is central to a molecular understanding of many biological self-assembly processes, including protein folding and the formation of lipid membranes (1-4). To understand the water-induced conformational changes in biopolymers that lead to functional structures, it is critical to study not only the effects of water on these molecules but also the reverse, i.e., the modified behavior of water in the vicinity of these molecules. To avoid the inherent chemical and structural complexities of present in biological molecules, it is instructive to consider model systems with tunable degrees of freedom. Arguably the simplest such model system is a smooth spherical particle immersed in a water bath (5-18). Such a rudimentary solute model captures two key factors in solvation, the size of the solute and the strength of its interactions with water.Small, methane-sized solutes are accommodated by water with only minor disruptions of the bulk hydrogen-bond network. The resulting small-solute hydration thermodynamics has been predicted successfully by approaches that take into account the molecular-scale density fluctuations in bulk water, including scaled-particle theory (5, 19) (see ref. 20 for a recent review), Pratt-Chandler theory (8), the Gaussian field model (21), an information theory model (22), and Lum-Chandler-Weeks (LCW) theory (23). With the addition of a large repulsive solute, it becomes impossible for water molecules to maintain their bulk hydrogen-bond structure (24), which also affects the wetting behavior. At ambient conditions the contact theorem predicts a near-zero water density at a flat hard wall (5). Based on this rigorous limit and an interpolation formula, Stillinger anticipated that...