STRUCTURE AND DOMAIN-DOMAIN INTERACTIONS OF THE GELATIN-BINDING SITE OF HUMAN 72-KIIODALTON TYPE-IV COLLAGENASE (GELATINASE-, MATRIX METALLOPROTEINASE-2)

We have shown previously that all three fibronectin type II modules of gelatinase A contribute to its gelatin affinity. In the present inves tigation we have studied the structure and module-module interactions of this gelatin-binding domain by circular dichroism spectroscopy and differential scanning calorimetry. Comparison of the T-m values of the thermal transitions of isolated type II modules with those of bimodul ar or trimodular proteins has shown that the second type II module is significantly more stable in the trimodular protein coll 123 (T-m = 54 degrees C) than in the single-module protein coll 2 (T-m = 44 degrees C) or in the bimodular proteins coll 23 (T-m = 47 degrees C) and coll 12 (T-m = 48 degrees C). Analysis of the enthalpy changes associated with thermal unfolding of the second type II module suggests that it i s stabilized by domain-domain interactions in coll 123. We propose tha t intimate contacts exist between the three tandem type II units and t hey form a single gelatin-binding site. Based on the three-dimensional structures of homologous metalloproteases and type II modules, a mode l is proposed in which the three type II units form an extension of th e substrate binding cleft of gelatinase A.