REFINEMENT OF THE NMR SOLUTION STRUCTURE OF THE GAMMA-CARBOXYGLUTAMICACID DOMAIN OF COAGULATION-FACTOR-IX USING MOLECULAR-DYNAMICS SIMULATION WITH INITIAL CA2+ POSITIONS DETERMINED BY A GENETIC ALGORITHM

A genetic algorithm (GA) successfully identified the calcium positions in the crystal structure of bovine prothrombin fragment 1 bound with calcium ions (bfl/Ca). The same protocol was then used to determine th e calcium positions in a closely related fragment, the Gla domain of c oagulation factor IX, the structure of which had previously been deter mined by NMR spectroscopy in the presence of calcium ions. The most fr equently occurring low-energy structure found by GA was used as the st arting structure for a molecular dynamics refinement. The molecular dy namics simulation was performed using explicit water and the Particle- Mesh Ewald method to accommodate the long-range electrostatic forces. While the overall conformation of the NMR structure was preserved, sig nificant refinement is apparent when comparing the simulation average structure with its NMR precursor in terms of the N-terminal (Tyr1-N) n etwork, the total number of hydrogen bonds, the calcium ion coordinati ons, and the compactness of the structure. It is likely that the place ment of calcium ions in the protein is critical for refinement. The ca lcium ions apparently induce structural changes during the course of t he simulation that result in a more compact structure.