Structural and Functional Studies of γ-Carboxyglutamic Acid Domains of Factor VIIa and Activated Protein C: Role of Magnesium at Physiological Calcium

Structural and Functional Studies of γ-Carboxyglutamic Acid Domains of Factor VIIa and Activated... Crystal structures of factor (F) VIIa/soluble tissue factor (TF), obtained under high Mg2+ (50mM Mg2+/5mM Ca2+), have three of seven Ca2+ sites in the γ-carboxyglutamic acid (Gla) domain replaced by Mg2+ at positions 1, 4, and 7. We now report structures under low Mg2+ (2.5mM Mg2+/5mM Ca2+) as well as under high Ca2+ (5mM Mg2+/45mM Ca2+). Under low Mg2+, four Ca2+ and three Mg2+ occupy the same positions as in high-Mg2+ structures. Conversely, under low Mg2+, reexamination of the structure of Gla domain of activated Protein C (APC) complexed with soluble endothelial Protein C receptor (sEPCR) has position 4 occupied by Ca2+ and positions 1 and 7 by Mg2+. Nonetheless, in direct binding experiments, Mg2+ replaced three Ca2+ sites in the unliganded Protein C or APC. Further, the high-Ca2+ condition was necessary to replace Mg4 in the FVIIa/soluble TF structure. In biological studies, Mg2+ enhanced phospholipid binding to FVIIa and APC at physiological Ca2+. Additionally, Mg2+ potentiated phospholipid-dependent activations of FIX and FX by FVIIa/TF and inactivation of activated factor V by APC. Since APC and FVIIa bind to sEPCR involving similar interactions, we conclude that under the low-Mg2+ condition, sEPCR binding to APC-Gla (or FVIIa-Gla) replaces Mg4 by Ca4 with an attendant conformational change in the Gla domain ω-loop. Moreover, since phospholipid and sEPCR bind to FVIIa or APC via the ω-loop, we predict that phospholipid binding also induces the functional Ca4 conformation in this loop. Cumulatively, the data illustrate that Mg2+ and Ca2+ act in concert to promote coagulation and anticoagulation. http://www.deepdyve.com/assets/images/DeepDyve-Logo-lg.png Journal of Molecular Biology Elsevier

Structural and Functional Studies of γ-Carboxyglutamic Acid Domains of Factor VIIa and Activated Protein C: Role of Magnesium at Physiological Calcium

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Publisher
Elsevier
Copyright
Copyright © 2013 Elsevier Ltd
ISSN
0022-2836
D.O.I.
10.1016/j.jmb.2013.02.017
Publisher site
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Abstract

Crystal structures of factor (F) VIIa/soluble tissue factor (TF), obtained under high Mg2+ (50mM Mg2+/5mM Ca2+), have three of seven Ca2+ sites in the γ-carboxyglutamic acid (Gla) domain replaced by Mg2+ at positions 1, 4, and 7. We now report structures under low Mg2+ (2.5mM Mg2+/5mM Ca2+) as well as under high Ca2+ (5mM Mg2+/45mM Ca2+). Under low Mg2+, four Ca2+ and three Mg2+ occupy the same positions as in high-Mg2+ structures. Conversely, under low Mg2+, reexamination of the structure of Gla domain of activated Protein C (APC) complexed with soluble endothelial Protein C receptor (sEPCR) has position 4 occupied by Ca2+ and positions 1 and 7 by Mg2+. Nonetheless, in direct binding experiments, Mg2+ replaced three Ca2+ sites in the unliganded Protein C or APC. Further, the high-Ca2+ condition was necessary to replace Mg4 in the FVIIa/soluble TF structure. In biological studies, Mg2+ enhanced phospholipid binding to FVIIa and APC at physiological Ca2+. Additionally, Mg2+ potentiated phospholipid-dependent activations of FIX and FX by FVIIa/TF and inactivation of activated factor V by APC. Since APC and FVIIa bind to sEPCR involving similar interactions, we conclude that under the low-Mg2+ condition, sEPCR binding to APC-Gla (or FVIIa-Gla) replaces Mg4 by Ca4 with an attendant conformational change in the Gla domain ω-loop. Moreover, since phospholipid and sEPCR bind to FVIIa or APC via the ω-loop, we predict that phospholipid binding also induces the functional Ca4 conformation in this loop. Cumulatively, the data illustrate that Mg2+ and Ca2+ act in concert to promote coagulation and anticoagulation.

Journal

Journal of Molecular BiologyElsevier

Published: Jun 12, 2013

References

  • Role of magnesium in factor XIa catalyzed activation of factor IX: calcium binding to factor IX under physiologic magnesium
    Agah, S.; Bajaj, S.P.
  • Contribution of magnesium in binding of factor IXa to the phospholipid surface: implications for vitamin K-dependent coagulation proteins
    Messer, A.S.; Velander, W.H.; Bajaj, S.P.
  • Mg(2+) binding to the Gla domain of factor X influences the interaction with tissue factor
    Persson, E.; Ostergaard, A.
  • Membrane association with multiple calcium ions: vitamin-K-dependent proteins, annexins and pentraxins
    Nelsestuen, G.L.; Ostrowski, B.G.
  • Nanoscale studies of protein-membrane interactions in blood clotting
    Morrissey, J.H.; Tajkhorshid, E.; Rienstra, C.M.
  • Recent estimates of the structure of the factor VIIa (FVIIa)/tissue factor (TF) and factor Xa (FXa) ternary complex
    Lee, C.J.; Chandrasekaran, V.; Wu, S.; Duke, R.E.; Pedersen, L.G.
  • Fibronectin-adherent monocytes express tissue factor and tissue factor pathway inhibitor whereas endotoxin-stimulated monocytes primarily express tissue factor: physiologic and pathologic implications
    Bajaj, M.S.; Ghosh, M.; Bajaj, S.P.
  • Antigenic characterization of endothelial cell-derived microparticles and their detection ex vivo
    Abid Hussein, M.N.; Meesters, E.W.; Osmanovic, N.; Romijn, F.P.; Nieuwland, R.; Sturk, A.
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    Emsley, P.; Lohkamp, B.; Scott, W.G.; Cowtan, K.

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