Open AccessThe role of exon 5 in fibroblast collagenase (MMP‐1) substrate specificity and inhibitor selectivity
Author(s) -
Knäuper Vera,
Patterson Margaret L.,
GomisRüth Franz X.,
Smith Bryan,
Lyons Alan,
Docherty Andrew J. P.,
Murphy Gillian
Publication year - 2001
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.2001.02062.x
Subject(s) - exon , chimera (genetics) , matrix metalloproteinase , collagenase , chemistry , microbiology and biotechnology , fibroblast , cleavage (geology) , biochemistry , enzyme , biology , gene , in vitro , paleontology , fracture (geology)
Interstitial collagen is degraded by members of the matrix metalloproteinase (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 chimera, consisting primarily of MMP‐1, with the region coded for by exon 5 replaced with the equivalent region of MMP‐3, a noncollagenolytic MMP. Unlike 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 cleavage of two synthetic substrates display an MMP‐3 phenotype, however, cleavage of gelatin is slightly impaired as compared to the parent enzymes. The K iapp values for the exon 5 chimera complexed with synthetic inhibitors and N‐terminal TIMP‐2 also show a more MMP‐3‐like behaviour. However, the k on values for N‐terminal TIMP‐1 and N‐terminal TIMP‐2 are more comparable to those for MMP‐1. These data show that the region of MMP‐1 coded for by exon 5 is involved in both substrate specificity and inhibitor selectivity and the structural basis for our findings is discussed.