Plasminogen binds to plasmin-modulated factor Xa by Ca2+- and C-terminal lysine-dependent and -independent interactions

Jean E. Grundy; Mark A. Hancock; Scott C. Meixner; Roger C. MacKenzie; Marlys L. Koschinsky; Edward L. G. Pryzdial

doi:10.1160/TH06-08-0476

Thrombosis and Haemostasis, Table of Contents

Thromb Haemost 2007; 97(01): 38-44
DOI: 10.1160/TH06-08-0476

Blood Coagulation, Fibrinolysis and Cellular Haemostasis

Schattauer GmbH

Plasminogen binds to plasmin-modulated factor Xa by Ca2+- and C-terminal lysine-dependent and -independent interactions

Jean E. Grundy

¹Canadian Blood Services, R&D Department, University of British Columbia/Centre for Blood Research, Department of Pathology and Laboratory Medicine, Vancouver, British Columbia, Canada

,

Mark A. Hancock

²Department of Biochemistry, Queen’s University, Kingston, Ontario, Canada

,

Scott C. Meixner

¹Canadian Blood Services, R&D Department, University of British Columbia/Centre for Blood Research, Department of Pathology and Laboratory Medicine, Vancouver, British Columbia, Canada

,

Roger C. MacKenzie

³Institute of Biological Sciences, National Research Council of Canada, Ottawa, Ontario, Canada

,

Marlys L. Koschinsky

²Department of Biochemistry, Queen’s University, Kingston, Ontario, Canada

,

Edward L. G. Pryzdial

¹Canadian Blood Services, R&D Department, University of British Columbia/Centre for Blood Research, Department of Pathology and Laboratory Medicine, Vancouver, British Columbia, Canada

› Author Affiliations

Abstract

Summary

Plasminogen binding to receptors involves both C-terminal lysine-dependent and -independent interactions. The latter are poorly understood. Our earlier work demonstrated a novel Ca²⁺-enhanced bivalent interaction between plasmin-cleaved FXa (FXa33/13) and plasminogen truncated at Lys78 (Lys-Pg). Here we hypothesized that the effects of Ca²⁺may enable dissection of the C-terminal lysine-dependent and -independent interactions. To evaluate the role of the Glu-plasminogen (Glu-Pg) amino acids 1–77, binding of FXa33/13 to immobilized Glu-Pg was compared to Lys-Pg by surface plasmon resonance. Under identical conditions, approximately half the amount of FXa33/13 bound to Glu-Pg. The simplest fit of data suggested a 2:1 plasminogen:FXa33/13 stoichiometry for both, which were proportionately enhanced by Ca²⁺. Only Lys-Pg demonstrated significant Ca²⁺-independent binding to FXa33/13. In the presence of Ca²⁺, weak C-terminal lysine-independent binding could be detected, but only for Glu-Pg. The elastase-generated plasminogen fragment encompassing the angiostatin-like kringle domains 1 to 3 (K1–3) inhibited binding of FXa33/13 to Lys-Pg, whereas fragments corresponding to kringle 4- and kringle 5-protease domain had no effect. Immobilized K1–3 binding to FXa33/13 had both Ca²⁺-dependent and -independent components. The principal KM_d for the interaction was 10-fold higher than Lys-Pg. In the presence of Ca²⁺, eACA inhibited FXa33/13 binding to K1–3 by 30%, but eliminated binding in the absence of Ca²⁺. These studies suggest that Ca²⁺-dependent and -independent binding of Lys-Pg to FXa33/13 are C-terminal lysine-dependent. The N-terminal 1–77 amino acids of Glu-Pg confer significant C-terminal lysine-independent binding, which may play a role during the initiating stages of plasminogen activation.

Keywords

Coagulation factors - protein function / activity - plasminogen (-fragments) - Gla-domain proteins

Full Text

References

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