Enthalpic Breakdown of Water Structure on Protein Active-Site Surfaces

Abstract

The principles underlying water reorganization around simple nonpolar solutes are well understood and provide the framework for the classical hydrophobic effect, whereby water molecules structure themselves around solutes so that they maintain favorable energetic contacts with both the solute and the other water molecules. However, for certain solute surface topographies, water molecules, due to their geometry and size, are unable to simultaneously maintain favorable energetic contacts with both the surface and neighboring water molecules. In this study, we analyze the solvation of ligand-binding sites for six structurally diverse proteins using hydration site analysis and measures of local water structure, in order to identify surfaces at which water molecules are unable to structure themselves in a way that maintains favorable enthalpy relative to bulk water. These surfaces are characterized by a high degree of enclosure, weak solute-water interactions, and surface constraints that induce unfavorable pair interactions between neighboring water molecules. Additionally, we find that the solvation of charged side chains in an active site generally results in favorable enthalpy but can also lead to pair interactions between neighboring water molecules that are significantly unfavorable relative to bulk water. We find that frustrated local structure can occur not only in apolar and weakly polar pockets, where overall enthalpy tends to be unfavorable, but also in charged pockets, where overall water enthalpy tends to be favorable. The characterization of local water structure in these terms may prove useful for evaluating the displacement of water from diverse protein active-site environments.

Figure 1

Water structure on simple and complex surfaces. (a) Water clathrate cage around a methane molecule. Optimal (b) and sub-optimal (c & d) water structure on a protein active site surface, illustrated using MD simulation snapshots of a part of streptavidin active-site. Red dashes in c & d indicate lost interactions that are maintained in optimal scenario.

Figure 2

Hydration of simple apolar surfaces. The most probable water configurations in investigated hydration sites on exterior surface of CB7 (top left) and aromatic surface of Phe side-chain (bottom left). Molecular surface of CB7 and Phe is shown, gray: carbon, red: oxygen and blue: nitrogen. Number density (middle) and probability distribution (right) of the interaction energy for water molecules on methane, CB7 and Phe surfaces, compared with the same distribution for pure water.

Figure 3

High energy hydration sites in apolar surface pockets. Top row: Abl 13 and Abl 16. Bottom row: FXa 21 and FXa 23. Left: Representative water configurations in labelled and neighboring hydration sites in the context of active-site molecular surface (gray: carbon, red: oxygen and blue: nitrogen). Right: Number density distribution of the interaction energy of hydration site water molecules with their first shell neighbors, compared with the same distribution for pure water.

Figure 4

Hydration of the amide side chain of Asn. Left: The most probable water configurations in hydration sites forming hydrogen bonds with the side-chain (only shown for Asn). Middle and Right: normalized probability distribution and number density distribution of the interaction energy of hydration site water molecules with their first shell neighbors, compared with the same distribution for pure water.

Figure 5

High energy hydration sites in polar surface pockets. The most probable water configurations of labelled and neighboring hydration sites and their interactions with the protein surface are shown for sites Abl 6 (left) and FXa 32 (middle). Number density distribution of the energy of hydration site water molecules with their first shell neighbors, compared with the same distribution for pure water (right).

Figure 6

Hydration of charged side-chains of Asp and Arg. Left and Middle: The most probable water configurations in hydration sites forming hydrogen bonds with the sidechain of Asp and Arg, respectively. Right: Number density distribution of the interaction energy of hydration site water molecules with their first shell neighbors, compared with the same distribution for pure water.

Figure 7

Hydration sites in charged surface pockets. The most probable water configurations of labelled and neighboring hydration sites and their interactions with the protein surface are shown for sites Casp 0 & 4 (left) and AChE 0 (middle). Number density distribution of the interaction energy of hydration site water molecules with their first shell neighbors, compared with the same distribution for pure water (right).

Figure 8

Displacement of hydration sites by ligand functional groups. Apolar ligand moieties displace FXa 21 & 23 (left) and Abl 6 (middle), while a carboxylate displaces sites Casp 0 and 4 (right).