Skip to Content
MilliporeSigma
  • In vitro reconstitution of complexes between pro-matrix metalloproteinase-9 and the proteoglycans serglycin and versican.

In vitro reconstitution of complexes between pro-matrix metalloproteinase-9 and the proteoglycans serglycin and versican.

The FEBS journal (2013-04-23)
Nabin Malla, Eli Berg, Achilleas D Theocharis, Gunbjørg Svineng, Lars Uhlin-Hansen, Jan-Olof Winberg
ABSTRACT

Previously, we have shown that a proportion of the matrix metalloproteinase-9 (MMP-9) synthesized by the macrophage cell line THP-1 binds to a chondroitin sulfate proteoglycan (CSPG) core protein to form a reduction-sensitive heteromer. It was also shown that the hemopexin-like (PEX) domain and the fibronectin-like (FnII) module in the enzyme are involved in heteromer formation. In this paper, we show that reduction-sensitive and SDS-stable heteromers may be reconstituted in vitro by mixing proMMP-9 with either serglycin, versican or CSPGs isolated from various monocytic cell lines. In addition, a strong but SDS-soluble proMMP-9·CSPG heteromer was formed. The two macromolecules in the SDS-stable reduction-sensitive heteromers were not linked together by disulfide bonds. As for the heteromer isolated from THP-1 cells, in vitro reconstituted SDS-stable and SDS-soluble heteromers showed weaker binding to gelatin than the proMMP-9 monomer. Furthermore, gelatin inhibited in vitro reconstitution of the heteromers, showing that the FnII module is involved in the complex formation. Tissue inhibitor of metalloproteinase (TIMP)-1 was not be detected in the proMMP-9·CSPG complexes. However, the presence of TIMP-1 inhibited formation of the SDS-soluble heteromer, but not the SDS-stable reduction-sensitive heteromer. This indicates that different regions in the PEX domain are involved formation of these heteromers.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Gelatin from cold water fish skin, solid
Sigma-Aldrich
Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 20% in H2O
Millipore
Gelatin from porcine skin, medium gel strength, suitable for microbiology
Millipore
Gelatin from porcine skin, suitable for microbiology, low gel strength
Millipore
Gelatin from porcine skin, suitable for microbiology, high gel strength
Sigma-Aldrich
Sodium dodecyl sulfate solution, BioUltra, for molecular biology, 10% in H2O
Sigma-Aldrich
Gelatin from bovine skin, Type B, powder, BioReagent, suitable for cell culture
Sigma-Aldrich
Gelatin from porcine skin, gel strength 80-120 g Bloom, Type A
Sigma-Aldrich
Gelatin from porcine skin, gel strength 300, Type A
Sigma-Aldrich
Gelatin from porcine skin, gel strength ~175 g Bloom, Type A
Sigma-Aldrich
Gelatin solution, Type B, 2% in H2O, tissue culture grade, BioReagent, suitable for cell culture
Sigma-Aldrich
Triton X-100, peroxide- and carbonyl-free
Sigma-Aldrich
Triton X-100, laboratory grade
Sigma-Aldrich
Triton X-45
Sigma-Aldrich
Triton X-100, BioXtra
Sigma-Aldrich
Prionex® Highly purified Type A, aqueous solution
Sigma-Aldrich
Triton X-100, for molecular biology
Sigma-Aldrich
Chondroitin sulfate A sodium salt from bovine trachea, lyophilized powder, BioReagent, suitable for cell culture
Millipore
Gelatin from porcine skin, suitable for microbiology, ultrahigh gel strength
Chondroitin sulfate sodium (marine), European Pharmacopoeia (EP) Reference Standard
Chondroitin sulfate sodium, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
Sodium dodecyl sulfate, ≥99.0% (GC), dust-free pellets
Sigma-Aldrich
Sodium dodecyl sulfate, ≥99.0%
Sigma-Aldrich
Sodium dodecyl sulfate, SAJ special grade, ≥97.0%
Sigma-Aldrich
Sodium dodecyl sulfate, ACS reagent, ≥99.0%
Cystine, European Pharmacopoeia (EP) Reference Standard
Octoxinol 10, European Pharmacopoeia (EP) Reference Standard
Sigma-Aldrich
Sodium dodecyl sulfate, 92.5-100.5% based on total alkyl sulfate content basis
Sigma-Aldrich
Sodium dodecyl sulfate, BioReagent, suitable for electrophoresis, for molecular biology, ≥98.5% (GC)
Sigma-Aldrich
Sodium dodecyl sulfate, ReagentPlus®, ≥98.5% (GC)