Refinement of the NMR solution structure of the gamma-carboxyglutamic acid 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 the calcium positions in a closely related fragment, the Gla domain of coagulation factor IX, the structure of which had previously been determined by NMR spectroscopy in the presence of calcium ions. The most frequently occurring low-energy structure found by GA was used as the starting structure for a molecular dynamics refinement. The molecular dynamics 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, significant refinement is apparent when comparing the simulation average structure with its NMR precursor in terms of the N-terminal (Tyr1-N) network, the total number of hydrogen bonds, the calcium ion coordinations, and the compactness of the structure. It is likely that the placement of calcium ions in the protein is critical for refinement. The calcium ions apparently induce structural changes during the course of the simulation that result in a more compact structure.