We propose to leverage recent advances in computer technology and algorithm development, together with the growing database of three-dimensional (3D) structures of Vitamin K-dependent (VKD) proteins and their complexes, to build reliable 3D complexes in electrically neutral solvent and with structural water molecules in place. It is our hypothesis that theoretical techniques are now at a level of sophistication and accuracy to warrant the judicious application to key coagulation systems: the prothrombin-prothombinase complex, which provides the penultimate step in the cascade (formation of thrombin) and the extrinsic tenase complex, which initiates the extrinsic blood coagulation cascade. We propose to provide solvent-equilibrated models for these systems. The plan:
AIM I : To model the prothrombinase-prothrombin complex, including the apparently flexible central fragment involving the kringle domains of prothrombin?the goal being to understand the molecular details of the multiple cleavages;
AIM : To complete a consensus model of TF/FVIIa/FXa followed by model studies of TFalone, TF as a dimer, and TF and its putative complexes with protein disulfide isomerase (PDI) (also to be modeled), glutathione and NO?the goal being to understand more completely the issues surrounding cryptic vs procoagulant TF;
AIM III : To complete a model study with solvation of the Vitamin K cycle reactions (enzymes: vitamin K carboxylase and vitamin K epoxide reductase) using quantum mechanics.
The final aim recognizes the need to not only develop all-atom, solvated, 3D structures, but also to provide insight into the quantum mechanical bond-breaking and bond-forming mechanisms that regulate coagulation. Underlying our applied work is the continual development of new techniques?modeling the interaction of protein domains to membrane surfaces( e.g. Gla domains to phospholipid surfaces rich in phosphatidylserine), improving protein-protein docking algorithms, modeling conformational changes in proteins (e.g. the conformational change of the C2 domain of Factor Va when binding to a membrane surface, FVIIa when binding TF, the Gla domain when binding calcium ions) and improving molecular dynamics methods. The developed complex structures will be made available through the internet and these will maintain value as a base for systematic improvement even as new experimental structures are solved.
The study of molecular processes in the blood coagulation cascade (here: prothrombin, factor Vila/Tissue factor, Vitamin K cycle) have high relevance to public health. These molecular processes have impact in many blood diseases, including hemophilia, stroke and plaque in the vascular system. Our research is aimed directly at understanding the molecular processes of normal and abnormal blood coagulation.
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