The matrix metalloptroteinases (MMPs) are a group of proteinases that are central to the degradation of extracellular matrix. They perform a variety of critical functions in the mammalian organism and in diseases such as arthritis periodontitis, atherosclerosis, and in tumor cell invasion and metastasis. Seventeen MMPs have been identified including four collagenases, two gelatinases, two stromelysins, four membrane-type MMPs (MT-MMPs), and five others. The principal investigator has for many years been making major contributions to our understanding of MMP function, and in this application he requests support to continue this in two areas. In the first specific aim the principal investigator proposes to define those aspects of collagenase structure that give it the ability to unwind and thus cleave the collagen triple helix. This will be done using recombinant technologies to generate chimeric or mutant enzymes. The interaction of MMP-1 and MMP-1/MMP-3 chimeras with synthetic triple helica peptides (supplied by collaborator Dr. Fields) that mimic sequences around the cleavage site of collagens with type I, II and III collagens will also be examined using fluorescent polarization and microcalorimetry techniques. In addition in collaboration with Dr. Bode the principal investigator aims to determine the crystal structure of catalytically inactive MMP-1 in complex wit one or more triple helical peptides. The second specific aim addresses the activation of proMMP 2 at the cell surface, a process that appears critical for cancer cell metastasis. A quantitative assay for proMMP 2 activation will be developed using radiolabelled catalytically inactive proMMP 2. The importance of cell attachment and the structure of the MT1-MMP molecule in the activation of MMP-will be examined by construction and cell surface expression of MMP-1/MT1-MMP chimeras containing separately the catalytic and C-terminal domains of MMP 1. The principal investigator also proposes to examine the role of cell surface TIMP-2 in this activation process.
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