title: Bivalency of plasminogen monoclonal antibodies is required for plasminogen bridging to fibrin and enhanced plasmin formation.
abstract: Binding of plasminogen to fibrin and cell surfaces is essential for fibrinolysis and pericellular proteolysis. We used surface plasmon resonance and enzyme kinetic analyses to study the effect of two mAbs (A10.2, CPL15) on plasminogen binding and activation at fibrin surfaces. A10.2 is directed against the lysine-binding site (LBS) of kringle 4, whereas CPL15 recognises a region in kringle 1 outside the LBS. In the presence of CPL15 and A10.2 mAbs, binding of plasminogen (K(d)=1.16+/-0.22 micromol/l) to fibrin was characterised by a mAb concentration-dependent bell-shaped isotherm. A progressive increase in the concentration of mAbs at the surface was also detected, and reached a plateau corresponding to the maximum of plasminogen bound. These data indicated that at low mAb concentration, bivalent plasminogen-mAb-plasminogen ternary complexes are formed, whereas at high mAb concentration, a progressive shift to monovalent plasminogen-mAb binary complexes is observed. Plasmin formation in the presence of mAbs followed a similar bell-shaped profile. Monovalent Fab fragments of mAb A10.2 showed no effect on the binding of plasminogen, confirming the notion that a bivalent mAb interaction is essential to increase plasminogen binding and activation at the surface of fibrin.
authors.affiliation: Plasminogen Activation in Cardiovascular Remodelling, Institut National de la Santé et de la Recherche Médicale, INSERM U460, UFR de Médecine Xavier Bichat, 16 rue Henri Huchard-BP 416, Paris, France.
authors.forename: Miguel
authors.initials: M
authors.lastname: Dominguez
authors.forename: Ramón
authors.initials: R
authors.lastname: Montes
authors.forename: José Antonio
authors.initials: JA
authors.lastname: Páramo
authors.forename: Eduardo
authors.initials: E
authors.lastname: Anglés-Cano
mesh_terms: D000911:Antibodies, Monoclonal / Q000276:immunology*; D001666:Binding Sites, Antibody; D002462:Cell Membrane / Q000378:metabolism; D005174:Factor XII / Q000378:metabolism; D005337:Fibrin / Q000378:metabolism*; D005341:Fibrinolysin / Q000378:metabolism*; D005342:Fibrinolysis; D005353:Fibronectins / Q000378:metabolism; D006801:Humans; D007140:Immunoglobulin Fab Fragments / Q000276:immunology; D007700:Kinetics; D008958:Models, Molecular; D010958:Plasminogen / Q000737:chemistry / Q000276:immunology* / Q000378:metabolism*; D011487:Protein Conformation; D020349:Surface Plasmon Resonance / Q000379:methods
chemical_list: D000911:Antibodies, Monoclonal; D005353:Fibronectins; D007140:Immunoglobulin Fab Fragments; D005174:Factor XII; D005337:Fibrin; D010958:Plasminogen; D005341:Fibrinolysin