Structure of Membrane-Bound Porcine Factor VIII

Abstract

Human Factor VIII (FVIII) is the co-factor of the serine protease Factor IXa (FIXa) in the membrane-bound tenase complex. Defects or deficiency of FVIII cause Hemophila A. Recombinant FVIII concentrate is the most effective cure for Hemophilia A. Porcine FVIII is highly homologous to human FVIII (84% sequence identity).In this work, we present for the first time, direct structural information for porcine FVIII, helically organized on lipid nanotubes (LNT) at closest to physiological conditions. This was achieved by combining direct structural data from Cryo-EM and molecular modeling. The 3D-structure at 15A resolution supports our hypothesis that the functional FVIII membrane-bound organization differs from that of the 3D-crystal structure in solution. Fitting the FVIII domains form the human crystal structure and the porcine homology model, within the 3D-Cryo-EM map, will allow us to define the inter- and intra-protein interfaces important for the FVIII function.Resolving the membrane-bound porcine FVIII structure by Cryo-EM will provide a scaffold for mapping functionally important FVIII interaction sites.A: 2D-Class average of porcine-FVIII, B: FFT of A. C: surface-representation of human-FVIII crystal-structure. The side-chains different with porcine-FVIII are shown in black. D: 3D-reconstruction of porcine-FVIII molecules onto LNT.View Large Image | View Hi-Res Image | Download PowerPoint Slide