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Measurement of Factor V Activity in Human Plasma Using a Microplate Coagulation Assay
Published on: September 9, 2012
Activated factor X cleaves factor VIII at arginine 562, limiting its cofactor efficiency.
J L Plantier1, V Rolli, C Ducasse
1Laboratoire d'hémobiologie EA4174, Faculté RTH Laennec, Université de Lyon, Université Lyon 1, Lyon, France. plantier@sante.univ-lyon1.fr
Journal of Thrombosis and Haemostasis : JTH
|October 31, 2009
Summary
Activated Factor X (FXa) cleaves Factor VIII (FVIII) at R562, a new inactivation pathway. This FXa-mediated FVIII degradation in the A2 domain impacts cofactor function and is inhibited by Von Willebrand factor and activated FIX.
Area of Science:
- Biochemistry
- Hematology
- Molecular Biology
Background:
- Factor VIII (FVIII) and its activated form (FVIIIa) are crucial for blood coagulation.
- Proteolysis by proteases like activated protein C (APC) and activated Factor X (FXa) regulates FVIII activity.
- APC cleaves FVIII at R562 in the A2 domain, fully inactivating it.
Purpose of the Study:
- To investigate the role of FXa cleavage at R562 in the A2 domain of FVIII.
- To understand the mechanism of FVIII inactivation by FXa.
Main Methods:
- Utilized an antibody (GMA012/R8B12) targeting the A2 C-terminus (A2C) to detect FXa activity.
- Generated an FVIII mutant at R562 (R562K) to assess the functional significance of this residue.
Main Results:
- FXa cleaves FVIII and FVIIIa within the A2 domain at R562, evidenced by A2C domain appearance.
- This cleavage requires phospholipids, occurs rapidly, and is inhibited by Von Willebrand factor and activated FIX.
- The R562K mutation enhanced FVIII activity and catalytic efficiency, preventing A2C fragment formation.
Conclusions:
- Identified a novel FXa-mediated FVIII degradation pathway involving A2 domain cleavage at R562.
- This mechanism contributes to the regulation of FVIII cofactor function.
- FXa-mediated cleavage at R562 is a significant factor in FVIII inactivation.
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