Nature adores a vacuum?

Well, ok, nature does not actually adore a vacuum, but what are we then to make of Bruce Berne's newest PNAS piece, in which he talks about water molecules in protein active sites that are so horribly unhappy that they would almost let a vacuum take their place?

The article comes on the heels of Friesner et al.'s comprehensive JMC 06 Glide XP article, which was an impressively comprehensive instance of how computational chemists can take into consideration the detailed physical chemistry of protein-ligand/drug interactions. As usual, electrostatics, van der Waals interactions, and desolvation among other things are taken care of. As usual, it turns out that in most earlier studies, the 'other half'-water-had not been considered. No wonder attempts to explain diabolically strong interactions like that of streptavidin-biotin (the strongest protein-ligand interaction known) resulted in drastic underestimates of the binding energy. With their latest framework, Berne et al. seek to remedy this situation. They make a couple of elementary sounding but oft neglected and important points:

1. Water is not going to be happy in a place in a protein where it is surrounded by a hydrophobic enclosure.

2. It is not going to be happy if it cannot make its full complement of hydrogen bonds with the protein/with other water molecules.

3. Even if it can make such bonds, it is not going to be happy if there are only one or two configurations in which it can do so.

4. Clearly, such water molecules are going to be more than happy to be kicked out by ligand atoms which can form their full complement of h-bonds with protein residues. These bonds, if they form part of a ring or if they are in close vicinity, have been called correlated hydrogen bonds. As per the authors' interpretation, its is these correlated hydrogen bonds that give special binding stability to examples like the streptavidin-biotin complex.

Obviously, like all life forms, water aspires to that highest ideal- freedom.

In the PNAS article, Berne et al. examine three cases, including streptavidin and COX 2 binding, in which water seems to be simply aching to get of the protein cavity. In the case of streptavidin, five water molecules form a metastable ring as depicted above. They do manage to form bonds with each other and with protein residues, but they are hopelessly trapped on top and bottom by hydrophobic parts. In this case, as explained in the JMC piece, the atom-atom pairwise addition principle does not work for hydrophobic enclosures. Clearly, the sum of the parts is not equal to the whole here, and the effect of two hydrophobic atoms, one each on top and bottom, is different from the calculated pairwise sum of effects for two atoms .

The word "metastable" is the best word I can think of for describing this unhappy situation for water. Surreal, simmering, patient, but waiting to become free.

In the case of COX 2, I saw a few statements which neither me nor my advisor could make sense of at first. Aristotle notwithstanding, it seems that there are actually some solvation situations which are almost as bad as a vacuum. Or even worse? Consider what the authors have to say:

The Cox-2 active site was found to contain no persistent hydration sites and is in fact entirely devoid of solvent in 80% of the simulation, despite the cavity sterically accommodating approximately seven water molecules. The high excess chemical potential of the binding-cavity solvent is due to an inability of the water molecules to make hydrogen bonds with the surrounding hydrophobic protein residues and other water molecules...An extreme case of hydrophobic enclosure is observed in the Cox-2 binding cavity, where no energetically stable solvent configurations appear to exist; insertion of ligand hydrophobic groups into such a region of persistent vacuum will result in substantially larger free-energy liberation than would be expected if the binding cavity were treated as solvated.

Well...I second the study; water, that humble but amazingly complex liquid, has often been neglected and underestimated in studies of protein-drug-ligand interactions.

Almost concomitantly, Essex et al. have published a report in JACS, in which they examined the nature of water molecules in active sites. They find that most of the water molecules are displaceable. Not surprisingly, they conclude that the binding affinity of the ligand depends upon the nature and environment of the water molecules. It's a good statistical study.

A central point that is coming to light through such studies is that the absolute hydrophobicity/hydrophilicity of a ligand certainly does not constitute the whole story. This may well be kept in mind by medicinal chemists who aim at modulating these properties of drug-like molecules from SAR studies. Sometimes, a single well-positioned ligand atom which displaces one or two extremely metastable water molecules can lead to orders of magnitude of binding affinity, which cannot be predicted from absolute characteristics of the ligand.

As usual, quality, not quantity, matters.

Addendum:
Another nice 3 page review I found in the rarely read Chemical Biology and Drug Design again makes the case. The review explores the thermodynamic aspects of drug-protein binding, and says among other things:

As the major contributors to the binding enthalpy are polar groups, a common misconception is that enthalpically driven compounds must be highly polar and that consequently their bioavailability will be compromised. In fact, what is often observed experimentally is that compounds with the same number of polar groups have vastly different binding enthalpies...To generate a favorable binding enthalpy, it is not the number of polar groups that matters but the quality of their interactions with the target. It is better to have few groups that establish strong interactions than a large number of groups mostly paying the desolvation penalty.

Again, by a "few groups", all that is needed is two functional groups that can kick out a metastable water molecule, and your job might be done.