Matrix metalloproteinase-1 takes advantage of the induced fit mechanism to cleave the triple-helical type I collagen molecule.

The collagenases are members of the matrix metalloproteinase family (MMP) that degrade native triple-helical type I collagen. To understand the mechanism by which these enzymes recognize and cleave this substrate, we studied the substrate specificity of a modified form of MMP-1 (FC) in which its active site region (amino acids 212-254) had been replaced with that of MMP-9 (amino acids 395-437). Although this substitution increased the activity of the enzyme toward gelatin and the peptide substrate Mca-PLGL(Dpa)AR-NH2 by approximately 3- and approximately 11-fold, respectively, it decreased the type I collagenolytic activity of the enzyme to 0.13%. The replacement of Gly233, the only amino acid in this region of FC that is conserved in all collagenase family members, with the corresponding Glu residue in MMP-9 resulted in a substantial decrease in the type I collagenolytic activity of the enzyme without affecting its general proteolytic activities. The kinetic parameters of the FC/G233E mutant for the collagen substrate were similar to those of the chimeric enzyme. In addition, substituting Gly233 for Glu in the chimera increased the collagenolytic activity of the enzyme by 12-fold. Interestingly, replacing Glu415 in MMP-9 with Gly, its corresponding residue in FC, endowed the enzyme with type I collagenolytic activity. The catalytic activity of the MMP-9 mutant toward triple-helical type I collagen was 2-fold higher than that of the collagenase chimera. These data in conjunction with the X-ray crystal structure of FC indicate that Gly233 provides the flexibility necessary for the enzyme active site to change conformation upon substrate binding. The flexibility provided by the Gly residue is essential for type I collagenolytic activity.