title: Assessment of Synthetic Matrix Metalloproteinase Inhibitors by Fluorogenic Substrate Assay.
abstract: Matrix metalloproteinases (MMPs) are a family of metzincin enzymes that act as the principal regulators and remodelers of the extracellular matrix (ECM). While MMPs are involved in many normal biological processes, unregulated MMP activity has been linked to many detrimental diseases, including cancer, neurodegenerative diseases, stroke, and cardiovascular disease. Developed as tools to investigate MMP function and as potential new therapeutics, matrix metalloproteinase inhibitors (MMPIs) have been designed, synthesized, and tested to regulate MMP activity. This chapter focuses on the use of enzyme kinetics to characterize inhibitors of MMPs. MMP activity is measured via fluorescence spectroscopy using a fluorogenic substrate that contains a 7-methoxycoumarin-4-acetic acid N-succinimidyl ester (Mca) fluorophore and a 2,4-dinitrophenyl (Dpa) quencher separated by a scissile bond. MMP inhibitor (MMPI) potency can be determined from the reduction in fluorescent intensity when compared to the absence of the inhibitor. This chapter describes a technique to characterize a variety of MMPs through enzyme inhibition assays.
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University, 95 Chieftan Way, Tallahassee, FL, 32306, USA.
authors.forename: Ty J
authors.initials: TJ
authors.lastname: Lively
authors.affiliation: Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA.
authors.forename: Dale B
authors.initials: DB
authors.lastname: Bosco
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University, 95 Chieftan Way, Tallahassee, FL, 32306, USA.
authors.forename: Zahraa I
authors.initials: ZI
authors.lastname: Khamis
authors.affiliation: Department of Chemistry and Biochemistry, Florida State University, 95 Chieftan Way, Tallahassee, FL, 32306, USA. qxsang@chem.fsu.edu.|Institute of Molecular Biophysics, Florida State University, 91 Chieftan Way, Tallahassee, FL, 32306, USA. qxsang@chem.fsu.edu.
authors.forename: Qing-Xiang Amy
authors.initials: QX
authors.lastname: Sang
mesh_terms: D002021:Buffers; D045744:Cell Line, Tumor; D031541:Fluorescence Resonance Energy Transfer / Q000379:methods*; D005456:Fluorescent Dyes / Q000737:chemistry*; D006801:Humans; D061965:Matrix Metalloproteinase Inhibitors / Q000494:pharmacology*; D020782:Matrix Metalloproteinases / Q000378:metabolism*
chemical_list: D002021:Buffers; D005456:Fluorescent Dyes; D061965:Matrix Metalloproteinase Inhibitors; D020782:Matrix Metalloproteinases
keywords: Fluorescence resonance energy transfer; Half maximal inhibitory concentration; Inhibition constant; Matrix metalloproteinase inhibitors; Matrix metalloproteinases