MMP9 Inhibitor Screening Assay Kit (Colorimetric) (ab139448)
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
MMP9 Inhibitor Screening Assay Kit (Colorimetric)
See all MMP9 kits -
Detection method
Colorimetric -
Sample type
Inhibitor compounds -
Assay type
Enzyme activity -
Product overview
Abcam MMP9 Inhibitor Screening Assay Kit (Colorimetric) (ab139448) is a complete assay system designed to screen MMP9 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 MMP9, 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 MMP9 Enzyme (Human, Recombinant) 1 x 48.5µl -
Research areas
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Function
May play an essential role in local proteolysis of the extracellular matrix and in leukocyte migration. Could play a role in bone osteoclastic resorption. Cleaves KiSS1 at a Gly-
-Leu bond. Cleaves type IV and type V collagen into large C-terminal three quarter fragments and shorter N-terminal one quarter fragments. Degrades fibronectin but not laminin or Pz-peptide. -
Tissue specificity
Produced by normal alveolar macrophages and granulocytes. -
Involvement in disease
Intervertebral disc disease
Metaphyseal anadysplasia 2 -
Sequence similarities
Belongs to the peptidase M10A family.
Contains 3 fibronectin type-II domains.
Contains 4 hemopexin repeats. -
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. -
Post-translational
modificationsProcessing of the precursor yields different active forms of 64, 67 and 82 kDa. Sequentially processing by MMP3 yields the 82 kDa matrix metalloproteinase-9.
N- and O-glycosylated. -
Cellular localization
Secreted, extracellular space, extracellular matrix. - Information by UniProt
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Alternative names
- 82 kDa matrix metalloproteinase-9
- 92 kDa gelatinase
- 92 kDa type IV collagenase
see all
Images
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Plot of OD vs time
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Inhibition of MMP9 by NNGHControl slope = 4.69E-03 OD/min
Inhibitor slope = 2.04E-04 OD/min
Inhibitor % activity remaining = (2.04E-04/4.69E-03) x 100 = 4.34%
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Inhibition of MMP9 by NNGH -
Example graph for Km and Vmax determinationKm=56.1 µM
Vmax=1.8 pmol/sec