The role of exon 5 in fibroblast collagenase (MMP-1) substrate specificityand inhibitor selectivity

Interstitial collagen is degraded by members of the matrix metalloproteinas e (MMP) family, including MMP-1. Previous work has shown that the region of MMP-1 coded for by exon 5 is implicated both in substrate specificity and inhibitor selectivity. We have constructed a chimeric enzyme, the exon 5 ch imera, consisting primarily of MMP-1, with the region coded for by exon 5 r eplaced with the equivalent region of MMP-3, a noncollagenolytic MMP. Unlik e MMP-3, the exon 5 chimera is capable of cleaving type I collagen, but the activity is only 2.2% of trypsin-activated MMP-1. 'Superactivation' of the chimera has no discernible effect, suggesting that the salt bridge formed in 'superactive' MMP-1 is not present. The kinetics for exon 5 chimera clea vage of two synthetic substrates display an MMP-3 phenotype, however, cleav age of gelatin is slightly impaired as compared to the parent enzymes. The K-iapp values for the exon 5 chimera complexed with synthetic inhibitors an d N-terminal TIMP-2 also show a more MMP-3-like behaviour. However, the k(o n) values for N-terminal TIMP-1 and N-terminal TIMP-2 are more comparable t o those for MMP-1. These data show that the region of MMP-1 coded for by ex on 5 is involved in both substrate specificity and inhibitor selectivity an d the structural basis for our findings is discussed.