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Abstract: Cryo-electron microscopy has the power to visualise lipid membranes at the closest to in vivo conditions. The structure of the lipid bilayer can be well resolved and the interactions between lipid–protein and protein–protein molecules followed at the molecular level. We undertook an extended Cryo-electron microscopy study to follow the factor VIII binding to phosphatidylserine containing lipid nanotubes at different lipid composition. Obtaining well ordered tubes is required to define the factor VIII membrane-bound structure. The observed alterations in the arrangement of the protein molecules are indicative for the flexibility of the membrane-bound factor VIII. Understanding the significance of these conformational changes is essential to comprehend the function of factor VIII in coagulation and as a drug for Hemophilia A. [Copyright &y& Elsevier]

Abstract: Factor VIII (FVIII) is a key protein in blood coagulation, deficiency or malfunction of which causes Haemophilia A. The sole cure for this condition is intravenous administration of FVIII, whose membrane-bound structure we have studied by Cryo-electron microscopy and image analysis. Self-assembled lipid nanotubes were optimised to bind FVIII at close to native conditions. The tubes diameter was constant at 30nm and the lipid bilayer resolved. The FVIII molecules were well defined, forming an 8.5nm thick outer layer, and appeared to reach the hydrophobic core of the bilayer. The two known FVIII atomic models were superimposed with the averaged 2D protein densities. The insertion of the FVIII within the membrane was evaluated, reaffirming that the membrane-binding C2 or C1–C2 domain(s) fully penetrate the outer leaflet of the lipid layer. The presented results lay the basis for new models of the FVIII overall orientation and membrane-binding mechanism. [Copyright &y& Elsevier]