Inactivation of plasminogen activator inhibitor-1 by specific proteolysis with stromelysin-1 (MMP-3).
Matrix metalloproteinase-3 (MMP-3 or stromelysin-1) specifically hydrolyzes the Ser(337)-Ser(338) (P10-P9) and Val(341)-Ile(342) (P6-P5) peptide bonds in human plasminogen activator inhibitor-1 (PAI-1). Cleavage is completely abolished in the presence of the metal chelators EDTA or 1,10-phenanthroline. A stabilized active PAI-1 variant was also cleaved by MMP-3. At an enzyme/substrate ratio of 1/10 at 37 degrees C, PAI-1 protein cleavage occurred with half-lives of 27 or 14 min for active or stable PAI-1 and was associated with rapid loss of inhibitory activity toward tissue-type plasminogen activator with half-lives of 15 or 13 min, respectively. A substrate-like variant of PAI-1, lacking inhibitory activity but with exposed reactive site loop, was cleaved with a half-life of 23 min, whereas latent PAI-1 in which a major part of the reactive site loop is inserted into the molecule, was resistant to cleavage. Biospecific interaction analysis indicated comparable binding of active, stable, and substrate PAI-1 to both proMMP-3 and MMP-3 (K(A) of 12-22 x 10(6) m(-1)), whereas binding of latent PAI-1 occurred with lower affinity (1.7-2.3 x 10(6) m(-1)). Stable PAI-1 bound to vitronectin was cleaved and inactivated by MMP-3 in a manner comparable with that of free PAI-1; however, the cleaved protein did not bind to vitronectin. Cleavage and inactivation of PAI-1 by MMP-3 may thus constitute a mechanism decreasing the antiproteolytic activity of PAI-1 and impairing the potential inhibitory effect of vitronectin-bound PAI-1 on cell adhesion and/or migration.
Lijnen
H R
HR
Center for Molecular and Vascular Biology and Laboratory for Pharmaceutical Biology and Phytopharmacology, University of Leuven, B-3000 Leuven, Belgium. roger.lijnen@med.kuleuven.ac.be
Arza
B
B
Van Hoef
B
B
Collen
D
D
Declerck
P J
PJ
D006801:Humans; D006868:Hydrolysis; D053507:Matrix Metalloproteinase 11; D008666:Metalloendopeptidases / Q000378:metabolism*; D008958:Models, Molecular; D017395:Plasminogen Activator Inhibitor 1 / Q000737:chemistry / Q000378:metabolism*; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000378:metabolism; D010959:Tissue Plasminogen Activator / Q000378:metabolism
D017395:Plasminogen Activator Inhibitor 1; D011994:Recombinant Proteins; D010959:Tissue Plasminogen Activator; D053507:Matrix Metalloproteinase 11; D008666:Metalloendopeptidases