Homology modeling of gelatinase catalytic domains and docking simulations of novel sulfonamide inhibitors

Three-dimensional models for the catalytic domain of gelatinases (MMP-9 and -2) have been constructed based on the X-ray crystal structure of MMP-3. Conformations of the loop segment which forms the bottom half of the S1' subsite but shows conformational diversity among the crystal structures of other MMPs have been explored by simulated annealing of each gelatinase model complexed with two highly potent "probe" inhibitors. Representative catalytic domain models have been selected for each gelatinase from the set of generated conformations based on shape complementarity of the loop to the probe inhibitors. The single model selected for MMP-9 was utilized to explain the structure-activity relationship of our novel sulfonamide inhibitors. Molecular dynamics (MD) simulations of the complex models revealed important features of the binding mechanism of our inhibitors: (i) the ligand carboxylate group coordinating to the catalytic zinc ion and hydrogen bonding to the Glu219 side chain, (ii) one of the sulfonyl oxygens forming hydrogen bonds with the main chain NHs (Leu181 and Ala182), (iii) the sulfonyl substituent making extensive hydrophobic contact with the S1' subsite. The gauche conformation exclusively adopted by the sulfonamide C-N-S-C torsion plays an important role in achieving the third binding feature by properly directing the substituent into the S1' subsite. Improvement of the inhibitory activity according to straight elongation of the sulfonyl substituent was attributed to an increase of the hydrophobic contact between the substituent and the S1' subsite. Structural modifications which alter the straight shape of the substituent lead to deterioration of the activity. On the other hand, the two candidate models selected for MMP-2 differ in the bottom shape of the S1' subsite: one with a channel-like subsite and the other with a pocket-like subsite resembling that of the MMP-9 model. The bottom shape was experimentally probed by chemical synthesis of inhibitors having elongated sulfonyl substituents whose terminal alkyl groups were shown by MD simulations to protrude from the S1' subsite bottom into the solvent. Gelatinase assays of these inhibitors showed that elongation of the substituent significantly reduces activity against MMP-9 while retaining activity against MMP-2, consequently increasing the selectivity between MMP-8 and -9. The results confirm that MMP-9 has a pocket-like S1' subsite with a floorboard and MMP-2 has a channel-like S1' subsite.