DIFFERENT COLLAGENASE GENE-PRODUCTS HAVE DIFFERENT ROLES IN DEGRADATION OF TYPE-I COLLAGEN

Vertebrate collagenases, matrix metalloproteinases (MMPs), cleave type I collagen at a single helical locus. We show here that rodent inters titial collagenases (MMP-13), but not human fibroblast collagenase (MM P-1), cleave type I collagen at an additional aminoitelopeptide locus. Collagenase cDNAs and chimeric constructs in pET-3d, juxtaposing MMP- 13 sequences amino-terminal to the active site in the catalytic domain and MMP-1 sequences carboxyl-terminal and vice versa, were expressed in Escherichia coli. Assays utilized collagen from wild type (+/+) mic e or mice that carry a targeted mutation (r/r) that encodes substituti ons in alpha 1(I) chains that prevent collagenase cleavage at the heli cal locus, MMP-13 and chimeric molecules that contained the MMP-13 seq uences amino-terminal to the active site cleaved (+/+) collagen at the helical locus and cleaved cross-linked (r/r) collagen in the aminotel opeptide (beta components converted to ct chains), Human MMP-1 and chi meric MMP-1/MMP-13 with MMP-1 sequences aminoterminal to the active si te cleaved collagen at the helical locus but not in the aminotelopepti de. All activities were inhibited by TIMP-1, 1,10-phenanthroline, and EDTA. Sequences in the distal two-thirds of the catalytic domain deter mine the aminotelopeptide-degrading capacity of MMP-13.