Role of Water in Protein Binding Interfaces

We could never expect to understand the chemistry of proteins completely without understanding aqueous solutions and the structure of water. – Walter Kauzmann   [1]
One of the goals in computational biochemistry is to accurately model molecular interactions to make better predictions for experimentalists to test. Since biological reactions occur in an aqueous solvent, understanding the role of water (and environment in general) has been central to progress in the field. Furthermore, water is intimately involved in both thermal and cold denaturation of proteins [see: Sai Janani Ganesan’s answer to How does cold denaturation of proteins happen? ] and hence a key player in understanding protein stability.
However, interactions between a protein molecule and its solvent is highly nuanced. The properties and fluctuations of hydration water molecules proximal to the protein surface (or interfacial waters) is vastly different from the bulk, and is known to affect protein motion [2] .  The relationship between this interfacial region and protein structure, dynamics and function is still very much an open question.
Understanding the role of water will be particularly valuable in binding studies  (and ergo drug design) due to the desolvation effect (removal of interacting water) that occurs when two protein molecules (or a protein and a ligand molecule) interact. Which consequently affects hydrogen bond strengths, salt bridges, electrostatic interactions, hydrophobic effect etc.
Figure 1: Binding of ligand changes hydrogen bond strengths (weak to strong) in a protein.
A recent study from Shoichet’s group demonstrates how water molecules in and around a cationic ligand binding site do not cause generic effects on binding [3] . The study compares electrostatic driven ligand binding to a buried cavity of a mutant Cytochrome c Peroxidase (CcP) protein, with an open cavity variant created by loop deletion. Ligands that are known to bind to the buried cavity were tested on the open cavity.  Ligands that (a) maintain ionic interactions with the site and form (b) a water interaction network were seen to have increased affinity to the open cavity.
Figure 2: Crystallographic pose for the ligand  3-fluorocatechol  in buried (gray) and open(pink) cavity. Ordered water in red. Ligand seen interacting favorably with water and protein.  [4]
While, ligands that could not favorably interact with both the waters and the site, changed orientation to preferentially interact with water. **
Figure 3: Crystallographic pose for the ligand 2-amino-5-methylthiazole  in buried (gray) and open(pink) cavity. Ordered water in red. Ligand  and protein seen interacting favorably with water.  [5]

Thus the role of water is complex and difficult to predict, and to adequately quantify these effects, structural, dynamical and thermodynamic information is necessary.  Structural effects are made more complex by the diversity in water-mediated interactions  [6] . In general, removing water from a binding site has  a thermodynamically favorable effect, due to an entropic gain when interfacial water is released to bulk solvent.  However, water mediated interaction can also lead to enthalpic gains resulting from new H-bonds. It is this fine balance that makes these interactions difficult to estimate, and more advances in both structural and thermodynamic characterization of such interactions is needed for improvements in drug design.

** As an MD-person, I think it might help to use MD before docking and after X-ray crystallography to make sure there are no artifacts in water interactions.


[1] Page on

[2] Slaving: Solvent fluctuations dominate protein dynamics and functions

[3] Roles for Ordered and Bulk Solvent in Ligand Recognition and Docking in Two Related Cavities

[4] Roles for Ordered and Bulk Solvent in Ligand Recognition and Docking in Two Related Cavities

[5] Roles for Ordered and Bulk Solvent in Ligand Recognition and Docking in Two Related Cavities

[6] Page on