Effect of species differences on stromelysin-1 (MMP-3) inhibitor potency -An explanation of inhibitor selectivity using homology modeling and chimeric proteins

For an animal model to predict a compound's potential for treating human di sease, inhibitor interactions with the cognate enzymes of separate species must be comparable. Rabbit and human isoforms of stromelysin-1 are highly h omologous, yet there are clear and significant compound-specific difference s in inhibitor potencies between these two enzymes. Using crystal structure s of discordant inhibitors complexed with the human enzyme, we generated a rabbit enzyme homology model that was used to identify two unmatched residu es near the active site that could explain the observed disparities, To tes t these observations, we designed and synthesized three chimeric mutants of the human enzyme containing the single (H224N and L226F) and double (H224N /L226F) mutations. A comparison of inhibitor potencies among the mutant and wild-type enzymes shows that the mutation of a single amino acid in the hu man enzyme, histidine 224 to asparagine, is sufficient to change the select ivity profile of the mutant to that of the rabbit isoform, These studies em phasize the importance of considering species differences, which can result from even minor protein sequence variations, for the critical enzymes in a n animal disease model. Homology modeling provides a tool to identify key d ifferences in isoforms that can significantly affect native enzyme activity .