Finding cryptic pockets computationally

The mobility of proteins is a constant source of wonder. I enjoy looking at experimentally-determined structures of small molecules bound to proteins, and it’s even more fun when the protein undergoes dramatic conformational changes to accommodate the ligand. But what may be fun for a chemist is a considerable challenge for molecular modeling: it’s hard enough to dock small molecules to a rigid model of a protein, and all the more so when an apparently flat protein surface yawns open to reveal a new pocket. In a recent paper in J. Comp. Chem., Olgun Guvench and collaborators at the University of New England College of Pharmacy and the University of Maryland look specifically for such “cryptic” binding sites.

The researchers used the cytokine IL-2, which is known to have cryptic pockets. In fact, small molecule inhibitors have been found that target the IL-2 receptor binding site in part by binding to cryptic pockets in the cytokine. The apo form of IL-2 (ie, without any small molecule bound) was used as a starting structure in a computational technique called Site Identification by Ligand Competitive Saturation (SILCS). In this approach, multiple molecular dynamic simulations are carried out with the protein “soaked” in a virtual solution of water and ligand (in this case very simple molecules such as benzene, propane, or acetonitrile). The idea is to let pockets form and see if the ligands bind in the pockets.

Molecular dynamics simulations, in which individual atoms within a protein are allowed to move, run the risk that the protein will deviate too far from a stable structure and denature completely. This can be avoided by introducing various restraints to keep atoms from moving too much, but if the restraints are too severe the protein is too rigid and you won’t see pockets form.

Also, as many people are painfully aware, small molecules can form aggregates in aqueous solution, and the same thing can happen in virtual water. In SILCS, the virtual fragments are programmed to repulse each other, keeping the fragments more or less distributed in solution.

The researchers found that they could in fact identify the cryptic pockets in IL-2, either by using relatively loose restraints or by running multiple unrestrained simulations and simply discarding those in which the protein denatured dramatically. Only the hydrophobic fragments found the cryptic binding sites, perhaps reflecting the relatively hydrophobic nature of the pockets. Additional pockets were also found, though whether these are real or not is unclear.

It’s still a long way from simulations with propane to running molecular dynamics screening simulations on hundreds or thousands of unique fragments, but given the increasing speed of processing power, perhaps the gap will be bridged sooner than expected.