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Biochemical characterization of the catalytic domain of membrane-type 4 matrix metalloproteinase

H Kolkenbrock1, L Essers, N Ulbrich

  • 1Deutsches Rheumaforschungszentrum Berlin, Germany.

Biological Chemistry
|October 30, 1999
PubMed

Insights

The catalytic domain of membrane-type 4 matrix metalloproteinase (MT4-MMP) is active but does not degrade extracellular matrix proteins. Tissue inhibitors of metalloproteinases (TIMPs) effectively inhibit MT4-MMP activity.

Area of Science:

  • Biochemistry
  • Enzymology
  • Molecular Biology

Background:

  • Matrix metalloproteinases (MMPs) are crucial enzymes involved in extracellular matrix (ECM) remodeling.
  • Membrane-type MMPs (MT-MMPs) play specific roles in cell adhesion, migration, and tissue development.
  • MT4-MMP (MMP-17) function and regulation are less understood compared to other MT-MMPs.

Purpose of the Study:

  • To characterize the catalytic activity and substrate specificity of the MT4-MMP catalytic domain.
  • To investigate the inhibitory effects of tissue inhibitors of metalloproteinases (TIMPs) on MT4-MMP activity.
  • To compare the functional properties of MT4-MMP with other MT-MMP family members.

Main Methods:

  • Expression and purification of the catalytic domain of MT4-MMP in Escherichia coli.
  • Tryptic activation of the recombinant proenzyme to generate the active catalytic domain.
  • Assays using fluorogenic substrates to determine catalytic activity.
  • Incubation with various ECM proteins (collagens, gelatin, fibronectin, laminin, decorin) to assess hydrolysis.
  • Inhibition studies using TIMP-1, TIMP-2, and a progelatinase A/TIMP-2 complex.

Main Results:

  • The recombinant catalytic domain of MT4-MMP was catalytically active towards a fluorogenic MMP substrate.
  • Unlike other MT-MMPs, MT4-MMP catalytic domain did not activate progelatinase A or degrade tested ECM proteins.
  • TIMP-1 effectively suppressed MT4-MMP activity.
  • TIMP-2 exhibited complex inhibitory behavior, with activity decreasing at low concentrations and increasing at high concentrations.
  • The progelatinase A/TIMP-2 complex also inhibited MT4-MMP, similar to TIMP-2 alone.

Conclusions:

  • The catalytic domain of MT4-MMP possesses enzymatic activity but lacks the broad substrate specificity and progelatinase A activation capabilities of other MT-MMPs.
  • MT4-MMP activity is effectively regulated by TIMP-1 and exhibits unique inhibition kinetics with TIMP-2.
  • These findings highlight distinct functional characteristics of MT4-MMP within the MT-MMP family, suggesting specialized roles in biological processes.

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