MMP9 Inhibitor Screening Assay Kit (Colorimetric) (ab139448)

Overview

  • 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.

  • 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.

  • Platform

    Microplate reader

Properties

  • 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
  • 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

    Defects in MMP9 may be a cause of susceptibility to intervertebral disc disease (IDD) [MIM:603932]; also known as lumbar disk herniation (LDH). IDD is one of the most common musculo-skeletal disorders and the predominant cause of low-back pain and unilateral leg pain.
    Defects in MMP9 are the cause of metaphyseal anadysplasia type 2 (MANDP2) [MIM:613073]. Metaphyseal anadysplasia consists of an abnormal bone development characterized by severe skeletal changes that, in contrast with the progressive course of most other skeletal dysplasias, resolve spontaneously with age. Clinical characteristics are evident from the first months of life and include slight shortness of stature and a mild varus deformity of the legs. Patients attain a normal stature in adolescence and show improvement or complete resolution of varus deformity of the legs and rhizomelic micromelia.
  • Sequence similarities

    Belongs to the peptidase M10A family.
    Contains 3 fibronectin type-II domains.
    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.
  • Post-translational
    modifications

    Processing 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
  • Alternative names

    • 82 kDa matrix metalloproteinase-9
    • 92 kDa gelatinase
    • 92 kDa type IV collagenase
    • Gelatinase B
    • GELB
    • MMP-9
    • MMP9
    • MMP9_HUMAN
    see all

Images

  • Control 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%

  • Km=56.1 µM

    Vmax=1.8 pmol/sec

Protocols

References

ab139448 has not yet been referenced specifically in any publications.

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