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A COMPARISON OF THE MEMBRANE BINDING PROPERTIES OF THE GAMMA-CARBOXY GLUTAMIC ACID PROTEINS OF BLOOD COAGULATION
Abstract number: P-S-055
Krisinger1 M.J., Pryzdial2 E.L.G., MacGillivray1 R.T.A.
11Department of Biochemistry and Molecular Biology 22Department of Pathology and Laboratory Medicine, University of British Columbia, Vancouver, Canada
How-to-cite Krisinger MJ, Pryzdial ELG, MacGillivray RTA. A COMPARISON OF THE MEMBRANE BINDING PROPERTIES OF THE GAMMA-CARBOXY GLUTAMIC ACID PROTEINS OF BLOOD COAGULATION. J Thromb Haemost 2007; 5 Supplement 2: P-S-055
Abstract
Introduction: The Gla-domain of vitamin K-dependent coagulation proteins facilitates essential membrane binding. However, a comparison of their membrane binding properties is not possible due to discrepancies between experimental conditions reported in previous studies. Using surface plasmon resonance (SPR), we have compared the affinity, kinetic and density binding parameters of Gla proteins (zymogens, activated enzymes and cofactors) under a single experimental condition.
Methods: Liposomes composed of 25% PS and 75% PC were stably immobilized to a biacore L1 chip. Protein interaction was monitored in 10 mM HEPES buffer, pH 7.4, containing 150 mM NaCl, 0.1% BSA and 5 mM CaCl2. The binding of Gla proteins was examined at multiple concentrations typically spanning a 10-fold range above and below the apparent equilibrium dissociation constant (appKd) of the interaction.
Results: The Gla protein-membrane interaction was not affected by mass transport, was Ca2+-dependent and completely reversible with EDTA. Non-specific binding was also negligible as des-Gla prothrombin showed absolutely no binding. Affinity varied by about 40-fold between the human Gla proteins analyzed, with the extremes in range represented by protein S (appKd = 73 2 nM) and protein C (appKd = 2.490.31 mM). A comparative examination of the kinetic binding profiles of the individual Gla proteins at an eqi-molar concentration (0.25 mM) revealed some striking differences in the overall shape of the kinetic curves reflecting the different kinetic on and off rate parameters intrinsic to individual Gla proteins.
Conclusions: We have shown that the Gla proteins of coagulation display a broad range of affinity, kinetic and density membrane binding properties. Differences in membrane binding properties also exist between specific zymogens and their respective activated enzymes. Since the Gla-domain sequence is unaltered in a zymogen-enzyme pair, a sequence independent effect is suggested to be the reason for these binding differences.
To cite this abstract use the following format:
Journal of Thrombosis and Haemostasis 2007; Volume 5, Supplement 2: abstract number
Session Details
| Date: |
01/08/2007
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| Time: |
00:00-00:00
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| Session name: |
XXIst ISTH Congress |
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| Location: |
Oxford, UK |
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