MMP-2 is a member of the family of matrix metalloproteinases (MMPs), which together cleave a broad range of tissue components. While this property of the MMPs is a beneficial feature of normal development and tissue adaptation, uncontrolled MMP-2 activity has been strongly associated with inflammatory diseases, such as periodontal disease and arthritis, and tumor expansion and metastasis. This application is designed to develop compounds, which specifically inhibit MMP-2 activity. Since cleavage of molecules by MMP-2 occurs only if there is binding between the enzyme and substrate molecules, the specific mechanism by which MMP-2 binds its main collagen substrates will be investigated. In a collaborative effort, molecular biology and protein structural analysis methods will be applied to first identify specific MMP-2 binding sites on collagen by screening a random peptide library and mapping the functional peptide sequences on collagen. To identify the precise collagen binding site residues on MMP-2, nuclear magnetic resonance studies will be used to analyze the MMP-2 collagen binding domain (CBD) complexed with synthetic peptides, which mimic the CBD binding sites on collagen. The specificity of the identified sites and amino acids will be tested in competitive ligand binding assays and by analyzing the effects of site-specific mutations in the CBD. Once the precise binding sites on both collagen and MMP-2 are defined, small molecules will be developed that can inhibit the full-length native MMP-2 activity by competing for substrate binding and by substituting binding site residues on the CBD. This will be accomplished in both MMP-2 ligand binding and activity assays, and in experiments with MMP-2 expressing cells. The proposed studies should define the specific binding site interactions between MMP-2 and its main collagen substrate and explore a new strategy to inhibit MMP-2 in inflammatory diseases and cancer based on competition for substrate binding.

Agency
National Institute of Health (NIH)
Institute
National Institute of Dental & Craniofacial Research (NIDCR)
Type
Research Project (R01)
Project #
5R01DE014236-03
Application #
6634719
Study Section
Oral Biology and Medicine Subcommittee 1 (OBM)
Program Officer
Shirazi, Yasaman
Project Start
2001-08-01
Project End
2006-05-31
Budget Start
2003-06-01
Budget End
2004-05-31
Support Year
3
Fiscal Year
2003
Total Cost
$325,125
Indirect Cost
Name
University of Texas Health Science Center San Antonio
Department
Dentistry
Type
Schools of Dentistry
DUNS #
800772162
City
San Antonio
State
TX
Country
United States
Zip Code
78229
Steffensen, Bjorn; Chen, Zhihua; Pal, Sanjay et al. (2011) Fragmentation of fibronectin by inherent autolytic and matrix metalloproteinase activities. Matrix Biol 30:34-42
Xu, Xiaoping; Mikhailova, Margarita; Ilangovan, Udayar et al. (2009) Nuclear magnetic resonance mapping and functional confirmation of the collagen binding sites of matrix metalloproteinase-2. Biochemistry 48:5822-31
Johnson, Dwight L; Carnes, David; Steffensen, Bjorn et al. (2009) Cellular effects of enamel matrix derivative are associated with different molecular weight fractions following separation by size-exclusion chromatography. J Periodontol 80:648-56
Stanley, Corey M; Wang, Yao; Pal, Sanjay et al. (2008) Fibronectin fragmentation is a feature of periodontal disease sites and diabetic foot and leg wounds and modifies cell behavior. J Periodontol 79:861-75
Murillo, Jesse; Wang, Yao; Xu, Xiaoping et al. (2008) Advanced glycation of type I collagen and fibronectin modifies periodontal cell behavior. J Periodontol 79:2190-9
Xu, Xiaoping; Chen, Zhihua; Wang, Yao et al. (2007) Inhibition of MMP-2 gelatinolysis by targeting exodomain-substrate interactions. Biochem J 406:147-55
Xu, Xiaoping; Chen, Zhihua; Wang, Yao et al. (2005) Functional basis for the overlap in ligand interactions and substrate specificities of matrix metalloproteinases-9 and -2. Biochem J 392:127-34
Xu, Xiaoping; Wang, Yao; Chen, Zhihua et al. (2005) Matrix metalloproteinase-2 contributes to cancer cell migration on collagen. Cancer Res 65:130-6
Xu, Xiaoping; Wang, Yao; Lauer-Fields, Janelle L et al. (2004) Contributions of the MMP-2 collagen binding domain to gelatin cleavage. Substrate binding via the collagen binding domain is required for hydrolysis of gelatin but not short peptides. Matrix Biol 23:171-81
Steffensen, Bjorn; Xu, Xiaoping; Martin, Pamela A et al. (2002) Human fibronectin and MMP-2 collagen binding domains compete for collagen binding sites and modify cellular activation of MMP-2. Matrix Biol 21:399-414