Figure 4

Comparison of the binding pocket entrance hydration sites as mapped using Szmap algorithm between the leishmanial L GSK-3 and L CRK3. Hydration sites with positive free energy as determined by Szmap computational analysis are depicted as red spheres in a ribbon representation of the protein. The hydration sites are connected with green lines depicting potential hydrogen bond networks that could be formed between them and neighboring residues. The depicted hydration sites potentially overlap with the bulky 3'-substituents of 6BIO analogues like those of compounds 11 and 17. Displacement and release of those water molecules by the compounds would favor binding affinity through the anticipated entropic gain. A) In L GSK-3, the indirubin substituent of position 3' occupies a region which overlaps with several unstable water molecules of the protein hydration shell. B) In L CRK3, the corresponding cluster of unstable water molecules is smaller and has a markedly lower overlap with the region occupied by the 3' substituent. As a result, binding affinity is not expected to be affected seriously by their displacement. In both figures, residues that differ between L GSK-3 and L CRK3 active sites are shown in stick representation and labelled (numbering based on L. donovani sequences). The ribbon fragments corresponding to the glycine-rich loop are colored green. The poses of analogues 11 and 17 are depicted as semi-transparent overlays on the 6BIO binding pose. For reasons of clarity and given that the poses of 11 and 17 are highly similar for each kinase, inhibitor 11 is depicted only in Figure 4A and inhibitor 17 in Figure 4B.