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. C onformations of the loop segment which forms the bottom half of the S1' sub site but shows conformational diversity among the crystal structures of oth er MMPs have been explored by simulated annealing of each gelatinase model complexed with two highly potent "probe" inhibitors. Representative catalyt ic domain models have been selected for each gelatinase from the set of gen erated conformations based on shape complementarity of the loop to the prob e inhibitors. The single model selected for MMP-9 was utilized to explain t he structure-activity relationship of our novel sulfonamide inhibitors. Mol ecular dynamics (MD) simulations of the complex models revealed important f eatures of the binding mechanism of our inhibitors: (i) the ligand carboxyl ate group coordinating to the catalytic zinc ion and hydrogen bonding to th e Glu219 side chain, (ii) one of the sulfonyl oxygens forming hydrogen bond s with the main chain NHs (Leu181 and Ala182), (iii) the sulfonyl substitue nt making extensive hydrophobic contact with the S1' subsite. The gauche co nformation 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 activi ty according to straight elongation of the sulfonyl substituent was attribu ted to an increase of the hydrophobic contact between the substituent and t he S1' subsite. Structural modifications which alter the straight shape of the substituent lead to deterioration of the activity. On the other hand, t he two candidate models selected for MMP-2 differ in the bottom shape of th e 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 exper imentally probed by chemical synthesis of inhibitors having elongated sulfo nyl substituents whose terminal alkyl groups were shown by MD simulations t o 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, con sequently increasing the selectivity between MMP-8 and -9. The results conf irm that MMP-9 has a pocket-like S1' subsite with a floorboard and MMP-2 ha s a channel-like S1' subsite.