Human tissue inhibitor of metalloproteinases 3 interacts with both the N- and C-terminal domains of gelatinases A and B. Regulation by polyanions.
We compared the association constants of tissue inhibitor of metalloproteinases (TIMP)-3 with various matrix metalloproteinases with those for TIMP-1 and TIMP-2 using a continuous assay. TIMP-3 behaved more like TIMP-2 than TIMP-1, showing rapid association with gelatinases A and B. Experiments with the N-terminal domain of gelatinase A, the isolated C-terminal domain, or an inactive progelatinase A mutant showed that the hemopexin domain of gelatinase A makes an important contribution to the interaction with TIMP-3. The exchange of portions of the gelatinase A hemopexin domain with that of stromelysin revealed that residues 568-631 of gelatinase A were required for rapid association with TIMP-3. The N-terminal domain of gelatinase B alone also showed slower association with TIMP-3, again implying significant C-domain interactions. The isolation of complexes between TIMP-3 and progelatinases A and B on gelatin-agarose demonstrated that TIMP-3 binds to both proenzymes. We analyzed the effect of various polyanions on the inhibitory activity of TIMP-3 in our soluble assay. The association rate was increased by dextran sulfate, heparin, and heparan sulfate, but not by dermatan sulfate or hyaluronic acid. Because TIMP-3 is sequestered in the extracellular matrix, the presence of certain heparan sulfate proteoglycans could enhance its inhibitory capacity.
Butler
G S
GS
School of Biological Sciences, University of East Anglia, Norwich, Norfolk NR4 7TJ, United Kingdom.
Apte
S S
SS
Willenbrock
F
F
Murphy
G
G
D000595:Amino Acid Sequence; D017364:Collagenases / Q000737:chemistry / Q000378:metabolism*; D005109:Extracellular Matrix / Q000378:metabolism; D018093:Gelatinases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D006493:Heparin / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D061965:Matrix Metalloproteinase Inhibitors; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000737:chemistry / Q000378:metabolism*; D009994:Osmolar Concentration; D000071228:Polyelectrolytes; D011108:Polymers / Q000378:metabolism*; D019717:Tissue Inhibitor of Metalloproteinase-3 / Q000378:metabolism*
D017364:Collagenases; D018093:Gelatinases; D006493:Heparin; D020778:Matrix Metalloproteinase 2; D020780:Matrix Metalloproteinase 9; D061965:Matrix Metalloproteinase Inhibitors; D008666:Metalloendopeptidases; D000071228:Polyelectrolytes; D011108:Polymers; D019717:Tissue Inhibitor of Metalloproteinase-3