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Structural changes in factor VIIa induced by Ca2+ and tissue factor studied using circular dichroism spectroscopy

P O Freskgård1, O H Olsen, E Persson

  • 1Vessel Wall Biology, Health Care Discovery, Novo Nordisk A/S, Gentofte, Denmark.

Protein Science : a Publication of the Protein Society
|August 1, 1996
PubMed
Summary

Calcium ions and tissue factor binding induce conformational changes in Factor VIIa (fVIIa), primarily affecting its Gla and catalytic domains. These changes are crucial for fVIIa

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Area of Science:

  • Biochemistry
  • Molecular Biology
  • Protein Structure

Background:

  • Factor VIIa (fVIIa) is a key enzyme in hemostasis, featuring distinct domains (Gla, EGF-like, protease) crucial for tissue factor (TF) interaction.
  • Calcium ions (Ca2+) are essential for fVIIa function, influencing its binding and catalytic activity, with most domains possessing Ca2+ binding sites.
  • Understanding Ca2+ and TF-induced conformational changes in fVIIa is vital for elucidating its activation mechanism.

Purpose of the Study:

  • To characterize and localize conformational changes in fVIIa upon binding of Ca2+ and TF using Circular Dichroism (CD) spectroscopy.
  • To investigate the role of specific fVIIa domains (Gla and protease) in these Ca2+-induced structural alterations.
  • To explore how TF binding impacts fVIIa's secondary structure and catalytic activity.

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Main Methods:

  • Circular Dichroism (CD) spectroscopy was employed to analyze the secondary and tertiary structural changes in intact fVIIa and its domain-deleted derivatives.
  • Comparative spectral analysis of intact fVIIa, Gla domain-deleted fVIIa, and protease domain-deleted fVIIa was performed.
  • Difference spectroscopy was used to model apo and Ca2+-bound forms of fVIIa and compare them to isolated Gla peptides.

Main Results:

  • Ca2+ binding induces ordering of alpha-helices in the Gla domain of fVIIa, consistent with models of other vitamin K-dependent proteins.
  • Tertiary structural alterations in the protease domain were observed upon Ca2+ binding, indicated by changes in the near-UV CD spectrum.
  • Formation of the fVIIa:TF complex resulted in secondary structural changes localized to the catalytic domain, suggesting a mechanism for TF-mediated activity enhancement.

Conclusions:

  • Ca2+ binding significantly impacts fVIIa structure, particularly ordering the Gla domain and altering the protease domain's tertiary structure.
  • TF binding induces specific secondary structural changes in the catalytic domain, likely contributing to the enhanced catalytic activity of fVIIa.
  • These findings provide insights into the allosteric regulation of fVIIa activity by Ca2+ and TF, crucial for blood coagulation.