title: Activity-Dependent Photoaffinity Labeling of Metalloproteases.
abstract: Metalloproteases, notably members of the matrix metalloprotease (MMP) and A Disintegrin And Metalloprotease (ADAM) families play crucial roles in tissue remodeling, the liberation of growth factors and cytokines from cell membranes (shedding) and cell-cell or cell-matrix interactions. Activity of MMPs or ADAMs must therefore be tightly controlled in time and space by activation of pro-enzymes upon appropriate stimuli and inhibition by endogenous tissue inhibitors of metalloproteases (TIMPs) or α2-macroglobulin to prevent irreversible tissue damage due to excessive degradation or uncontrolled release of potent inflammatory mediators, such as tumor necrosis factor-α (TNF-α).Although there is a wide range of methods to measure the amount of metalloproteases based on immunological approaches, relatively little is known about the activation status of a given enzyme at any given time and location. This information is, however, critical in order to understand the function and possible implication of these enzymes in disease. Since metalloproteases use an active-site bound water molecule to cleave the peptide bond, it is not possible to apply known active-site-directed labeling approaches with electrophilic "warheads." We therefore developed novel metalloprotease inhibitors that contain a photoactivatable trifluoromethylphenyldiazirine group and show that such inhibitors are suitable for activity-dependent photoaffinity labeling of MMPs and ADAMs.
authors: Department of Pharmacy, Analytical Biochemistry, University of Groningen, Antonius Deusinglaan 1, 9713, AV, Groningen, The Netherlands.
authors: Laurette
authors: L
authors: Prely
authors: Department of Pharmacy, Analytical Biochemistry, University of Groningen, Antonius Deusinglaan 1, 9713, AV, Groningen, The Netherlands.
authors: Theo
authors: T
authors: Klein
authors: Leiden Institute of Chemistry and the Netherlands Proteomics Center, Leiden University, Einsteinweg 55, 2300, RA, Leiden, The Netherlands.
authors: Paul P
authors: PP
authors: Geurink
authors: Department of Pharmacy, Analytical Biochemistry, University of Groningen, Antonius Deusinglaan 1, 9713, AV, Groningen, The Netherlands.
authors: Krisztina
authors: K
authors: Paal
authors: Bio-Organic Synthesis Groups, Leiden Institute of Chemistry, Leiden University, Einsteinweg 55, 2300, RA, Leiden, The Netherlands.
authors: Herman S
authors: HS
authors: Overkleeft
authors: Department of Pharmacy, Analytical Biochemistry, University of Groningen, Antonius Deusinglaan 1, 9713, AV, Groningen, The Netherlands. r.p.h.bischoff@rug.nl.
authors: Rainer
authors: R
authors: Bischoff
mesh_terms: D000818:Animals; D045744:Cell Line, Tumor; D003470:Culture Media; D004789:Enzyme Activation; D006801:Humans; D020128:Inhibitory Concentration 50; D045726:Metalloproteases / Q000378:metabolism*; D051379:Mice; D019920:Photoaffinity Labels; D019715:Tissue Inhibitor of Metalloproteinase-1 / Q000378:metabolism; D014409:Tumor Necrosis Factor-alpha / Q000378:metabolism
chemical_list: D003470:Culture Media; D019920:Photoaffinity Labels; C560822:TIMP1 protein, human; D019715:Tissue Inhibitor of Metalloproteinase-1; D014409:Tumor Necrosis Factor-alpha; D045726:Metalloproteases
keywords: A Disintegrin And Metalloprotease (ADAM); Matrix metalloprotease (MMP); Metalloprotease; Photoaffinity labeling; Trifluoromethylphenyldiazirine