Abstract

Formation of the intrinsic tenase-complex, is an essential event in the procoagulant reactions that lead to clot formation. The tenase-complex is formed when the activated serine protease, factor IXa (FIXa), and its cofactor factor VIIIa (FVIIIa) assemble on the phospholipid surface of activated platelets in the presence of calcium ions to proteolytically convert the zymogen factor X (FX) into its active form FXa. The physiological relevance of the tenase complex is evident in hemophilia A or B patients, who present with bleeding disorders due to quantitative and/or qualitative deficiencies of FVIII or FIX. In this work we present preliminary models of the FVIIIa-FIXa-FX complex that were generated based on the published X-ray structures of FIXa (

PNAS
1995
;
92
:
9796
–800
.), factor Va (
PNAS
2004
;
101
:
8918
–23
.), factor Xa (
PNAS
1995
;
95
:
6630
–5
.) and the FVIII homology model (
Blood
2002
;
99
:
1215
–23
.). In a first step we developed new, refined, theoretical models of FVIIIa and FX zymogen via homology modeling, inter-domain docking/simulation and new loop simulation algorithms. This was followed by Pseudo-Brownian protein-protein docking in internal coordinates between FVIIIa and FIXa and between FIXa and FX with internal coordinate mechanics (
Proteins
2003
;
52
:
113
–7
.). Flexibility of the molecules was partially accounted for by the use of a soft interaction energy function precalculated on a grid. Several models of the tenase-complex were selected based on structural criteria and agreement with known experimental data such as site directed mutagenesis, naturally occurring FVIII and FIX mutations reported in hemophilia, and overall location of the membrane plane with regard to the enzyme active site and peptide studies. These 3D models of the tenase-complex will be useful tools to guide future site directed mutagenesis and drug design studies aimed at improving functionality and half-life of FVIII and FIX and will contribute a better understanding of the role of the tenase-complex in the blood coagulation cascade.

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