Plasminogen binding properties of macrophage inflammatory protein (MIP)-2alpha.
The chemokine macrophage inflammatory protein (MIP)-2alpha was identified as a plasminogen binding protein by phage display analysis. MIP-2alpha and a truncated form lacking 5 lysine residues in the COOH-terminal region (mut-MIP-2alpha) were expressed in E. coli and purified to apparent homogeneity. Purified MIP-2alpha but not mut-MIP-2alpha bound specifically to plasminogen, with K(A) of 3.7 X 10(5) M(-1) for the interaction of plasminogen with surface-bound MIP-2alpha. Binding and competition experiments indicated that the interaction involves the region comprising the first 3 kringles of plasminogen and the COOH-terminal lysine-rich domain of MIP-2alpha. Activation of plasminogen bound to surface-associated MIP-2alpha by two-chain urokinase-type plasminogen activator (tcu-PA) was about 2.5-fold more efficient than in solution (catalytic efficiency k(cat)K(M) of 0.1 microM(-1)s(-1), as compared to 0.04 microM(-1)s(-1). In contrast, binding of plasminogen to MIP-2alpha in solution was very weak, as evidenced by the absence of competition of MIP-2alpha with lysine-Sepharose or with human THP-1 cells for binding of plasminogen. In agreement with this finding, addition of excess MIP-2alpha did not affect the main functional properties of plasmin(ogen) in solution, as indicated by unaltered activation rates of plasminogen by tcu-PA or tissue-type plasminogen activator (t-PA), t-PA-mediated fibrinolysis, and inhibition rate of plasmin by alpha2-antiplasmin. Thus, association of MIP-2alpha with surfaces exposes its COOH-terminal plasminogen-binding site, and may result in enhanced local plasmin generation.
Institut de Recerca Oncològica, Hospital Duran i Reynals, Barcelona, Spain.
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Arza
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Félez
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Fábregas
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Laroche
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Collen
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Lijnen
D001665:Binding Sites; D054426:Chemokine CXCL2; D019743:Chemokines, CXC; D004730:Endothelium, Vascular / Q000166:cytology; D004789:Enzyme Activation / Q000187:drug effects; D005342:Fibrinolysis / Q000187:drug effects; D006801:Humans; D036341:Intercellular Signaling Peptides and Proteins; D007700:Kinetics; D018082:Kringles; D007948:Leukemia, Monocytic, Acute / Q000473:pathology; D008239:Lysine / Q000378:metabolism; D008264:Macrophages / Q000378:metabolism; D015846:Monokines / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D010446:Peptide Fragments / Q000378:metabolism; D010958:Plasminogen / Q000378:metabolism*; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000378:metabolism; D017384:Sequence Deletion; D010959:Tissue Plasminogen Activator / Q000494:pharmacology; D014407:Tumor Cells, Cultured; D014568:Urokinase-Type Plasminogen Activator / Q000494:pharmacology
C499706:CXCL2 protein, human; D054426:Chemokine CXCL2; D019743:Chemokines, CXC; D036341:Intercellular Signaling Peptides and Proteins; D015846:Monokines; D010446:Peptide Fragments; D011993:Recombinant Fusion Proteins; D010958:Plasminogen; D010959:Tissue Plasminogen Activator; D014568:Urokinase-Type Plasminogen Activator; D008239:Lysine