Binding of human factor VIII to phospholipid vesicles

• Published in: The Journal of biological chemistry 1990-01, Vol.265 (2), p.815-822
• Main Authors: GILBERT, G. E, FURIE, B. C, FURIE, B
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; blood coagulation; Blood coagulation. Blood cells; Chromatography, Gel; Coagulation factors; Dansyl Compounds - metabolism; Energy Transfer; Factor VIII - metabolism; Fundamental and applied biological sciences. Psychology; Humans; Molecular and cellular biology; Phosphatidylcholines - metabolism; Phosphatidylserines - metabolism; Phospholipids - metabolism; Spectrometry, Fluorescence; vesicles
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1016/s0021-9258(19)40122-1

Factor VIII, a protein cofactor involved in blood coagulation, functions in vitro on a phospholipid membrane surface to greatly increase the rate of factor X activation by factor IXa. Using gel filtration, rapid sedimentation, and resonance energy transfer we have studied the interaction of recombinant-derived human factor VIII with small and large unilamellar phospholipid vesicles composed of phosphatidylserine and phosphatidylcholine. Resonance energy transfer, from intrinsic fluorophores in factor VIII to dansyl-phosphatidylethanolamine incorporated into vesicles, has been adapted for quantitative equilibrium measurements. Factor VIII binds rapidly and reversibly to small and large vesicles. At 8 degrees C the interaction of factor VIII with small vesicles fits a simple bimolecular model with a KD of 2 nM and a phospholipid binding site defined by 180 phospholipid monomers. At 25 degrees C the binding of factor VIII to small vesicles containing 20% phosphatidylserine can be described by an apparent KD of 4 nM; the phospholipid/protein ratio at saturation was 170. Binding to large vesicles was demonstrated with a KD of 2 nM and a phospholipid/protein ratio at saturation of 385. Binding was dependent upon the phosphatidylserine mole fraction and was nonlinear from 0 to 30% phosphatidylserine content. A direct comparison of factor VIII and factor V binding indicated that the affinity of factor V to phospholipid vesicles was equivalent to that of factor VIII and that the phosphatidylserine requirement was lower. A model is proposed to explain the nonlinear phosphatidylserine dependence of binding for factor VIII.