MMP12 Inhibitor Screening Assay Kit (Colorimetric) (ab139441)
Key features and details
- Assay type: Enzyme activity
- Detection method: Colorimetric
- Platform: Microplate reader
- Sample type: Inhibitor compounds
Overview
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Product name
MMP12 Inhibitor Screening Assay Kit (Colorimetric)
See all MMP12 kits -
Detection method
Colorimetric -
Sample type
Inhibitor compounds -
Assay type
Enzyme activity -
Product overview
Abcam's MMP12 Inhibitor Screening Assay Kit (Colorimetric) (ab139441) is a complete assay system designed to screen MMP12 inhibitors using a thiopeptide as a chromogenic substrate (Ac-PLG-[2-mercapto-4-methyl-pentanoyl]-LG-OC2H5). The MMP cleavage site peptide bond is replaced by a thioester bond in the thiopeptide. Hydrolysis of this bond by an MMP produces a sulfhydryl group, which reacts with DTNB [5,5’-dithiobis(2-nitrobenzoic acid), Ellman’s reagent] to form 2-nitro-5-thiobenzoic acid, which can be detected by its absorbance at 412 nm (e=13,600 M-1cm-1 at pH 6.0 and above). The assays are performed in a convenient 96-well microplate format.
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Notes
This kit is useful to screen inhibitors of MMP12, a potential therapeutic target. The MMP inhibitor NNGH is also included as a prototypic control inhibitor.
Thiol inhibitors should not be used with this kit, as they may interfere with the colorimetric assay.
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Platform
Microplate reader
Properties
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Storage instructions
Please refer to protocols. -
Components 1 x 96 tests 96-well Clear Microplate (1/2 Volume) 1 unit Colorimetric Assay Buffer 1 x 20ml MMP Inhibitor 1 x 50µl MMP Substrate 1 x 50µl MMP12 Enzyme (Human, Recombinant) 1 x 14µl -
Research areas
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Function
May be involved in tissue injury and remodeling. Has significant elastolytic activity. Can accept large and small amino acids at the P1' site, but has a preference for leucine. Aromatic or hydrophobic residues are preferred at the P1 site, with small hydrophobic residues (preferably alanine) occupying P3. -
Tissue specificity
Found in alveolar macrophages but not in peripheral blood monocytes. -
Sequence similarities
Belongs to the peptidase M10A family.
Contains 4 hemopexin-like domains. -
Domain
The conserved cysteine present in the cysteine-switch motif binds the catalytic zinc ion, thus inhibiting the enzyme. The dissociation of the cysteine from the zinc ion upon the activation-peptide release activates the enzyme. -
Cellular localization
Secreted > extracellular space > extracellular matrix. - Information by UniProt
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Alternative names
- EC 3.4.24.65
- HME
- Macrophage elastase
see all
Images
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Plot of OD vs. time.
Slope=V=4.85E-03 OD/min.
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Inhibition of MMP12 by NNGH (100 nM). Example of inhibitor data.control slope = 5.08E-03 OD/min
inhibitor slope = 4.82E-04 OD/min
inhibitor % activity remaining = (4.82E-04/5.08E-03) x 100 = 9.49% -
Example graph for Km and Vmax determinationActivity of a control sample =
(4.85E-03OD/min x 1E-04L)/(13,600M-1cm-1 x 0.5cm)=
7.13E-11 mol/min at 37°C, 100?M thiopeptolide -
Inhibition of MMP12 by NNGH (100 nM).
