Inactivation of the serpin alpha(2)-antiplasmin by stromelysin-1.
Matrix metalloproteinase-3 (MMP-3 or stromelysin-1) hydrolyzes the Met(374)-Ser(375) (P3-P2), Glu(416)-Leu(417) and Ser(432)-Leu(433) peptide bonds in human alpha(2)-antiplasmin (alpha(2)-AP), the main physiological plasmin inhibitor. Cleavage is completely abolished in the presence of the MMP inhibitors EDTA or 1,10-phenanthroline. At enzyme/substrate ratio of 1:10 at 37 degrees C, alpha(2)-AP protein cleavage occurs with a half-life of 8 min, and is associated with rapid loss of inhibitory activity towards plasmin with a half-life of 5 min. alpha(2)-AP cleaved by MMP-3 does no longer form a stable complex with plasmin, as shown by SDS-PAGE, and does no longer interact with plasminogen, as shown by crossed immunoelectrophoresis with plasminogen added to the gel. These data are compatible with the removal of a COOH-terminal fragment containing the reactive site peptide bond and the plasmin(ogen)-binding site. In addition, MMP-3 cleaves the Pro(19)-Leu(20) peptide bond in alpha(2)-AP, thereby removing the fibrin-binding site from the inhibitor. A dysfunctional alpha(2)-AP variant (Ala-alpha(2)-AP or alpha(2)-AP Enschede), with an alanine insertion in the reactive site sequence converting it from a plasmin inhibitor into a substrate, was also efficiently cleaved by MMP-3 (half-life of 13 min at 37 degrees C and enzyme/substrate ratio of 1:10). Cleavage and inactivation of alpha(2)-AP by MMP-3 may constitute a mechanism favoring local plasmin-mediated proteolysis.
Lijnen
H R
HR
Center for Molecular and Vascular Biology, University of Louvain, Campus Gasthuisberg, O&N, Herestraat 49, B-3000, Leuven, Belgium. roger.lijnen@med.kuleuven.ac.be
Van Hoef
B
B
Collen
D
D
D003720:Densitometry; D004591:Electrophoresis, Polyacrylamide Gel; D005341:Fibrinolysin / Q000737:chemistry; D007123:Immunoelectrophoresis, Two-Dimensional; D019278:Matrix Metalloproteinase 3 / Q000737:chemistry* / Q000494:pharmacology; D010446:Peptide Fragments / Q000737:chemistry; D013997:Time Factors; D000979:alpha-2-Antiplasmin / Q000037:antagonists & inhibitors / Q000737:chemistry*
D010446:Peptide Fragments; D000979:alpha-2-Antiplasmin; D005341:Fibrinolysin; D019278:Matrix Metalloproteinase 3