Simulation of conformational changes and coordinating dynamics in molecular motors

The aims of this thesis are to simulate the dynamics of F1-ATPase and related molecular motors and to investigate their conformational changes and coordinating mechanisms. The structure and functionality of F1-ATPase and other molecular motors were explored with the aid of structural analysis and molecular simulation, respectively. Molecular dynamics was employed to study the dynamics of F1-ATPase and other molecular motors. Steered molecular dynamics was used to explore the unbinding pathways of ATP from F1-ATPase. Structural properties of molecular motors retrieved from the Protein Data Bank were studied in tandem with the simulation results. The structural fluctuations of ADP/Mg:ADP and ATP/Mg:ATP were studied in three different molecular systems. Periodic boundary conditions and spherical boundary conditions were used for comparison reasons. Structural parameters such as inter atomic distances, bonds, angles and dihedral angles were obtained by data mining of crystal structures of molecular motors and from simulation results. The coordination dynamics of Mg2+ in molecular motors was investigated. Alignments of ADP, Mg:ADP, ATP and Mg:ATP were done to explore their flexibility and conformational diversity in molecular motors. The role of water molecules in determining the conformational fluctuations of ADP and ATP was investigated. The functional role played by Mg2+ was correlated with its coordination diversity. The pathways of ATP nucleotide upon unbinding were analysed and the conformational changes triggered in ab subunits by this unbinding process are presented. These changes were studied using structural alignments and Ramachandran plots. The dynamics of hydrogen bonds during unbinding was determined by measuring the interatomic distances between an electronegative atom and a hydrogen atom bonded to oxygen or nitrogen. The simulation results show agreement with the experimental results obtained by X-ray diffraction or NMR. The alignment analysis showed that the tight pocket of molecular motors diminishes the flexibility of ADP/ATP while the presence of water allows for higher flexibilities. The extensibility of the ADP and ATP molecule was shown using radius of gyration to be higher in the loose pockets and smaller in tight pockets of molecular motors. The diversity of conformations adopted by the phosphate region of ATP in crystal has been reported previously for a small number of ATP molecules. The main cause of this conformational diversity is the presence of Mg2+. As it was shown, the presence of Mg2+ is likely to facilitate the structural changes of the ribose base ring. This in turn drives the phosphate group to adapt different conformational states corresponding to ribose structural arrangements. These structural changes in ribose also cause the adenine to adopt different conformational states. The investigations of RMSD, RMSF and radius of gyration of ADP and ATP in the systems investigated were used to assess their structural extensibility and stability. These results together with the interaction energy investigation between water molecules or protein and ATP suggest that ATP is structurally much more unstable when coordinated by Mg2+ and the number of water molecules in pocket is high. ATP has a more stable behavior in the proteins where the number of water molecules is significantly lower. ATP appears to have the highest degree of structural instability in solvent followed by loose pocket and is most stable in tight pocket. Simulation results and data mining investigations provided a detailed picture of conformational fluctuations of ADP/ATP and Mg2+ coordinating dynamics in molecular motors. The agreement between simulation and experimental data allowed exploring new mechanisms as those that take place during unbinding of ATP from F1-ATPase. SMD investigations show that Mg2+ remains in the pocket after ATP is unbound. The hydrogen bonds were shown to form and break during the unbinding process. They are closely related with the force required to unbind ATP from F1-ATPase. Investigations of water density in the hydratation shells and pRDF show that in the tight pocket the number of water molecules is small. In loose or open pocket water molecules are in a bigger number determining ADP/ATP to adopt more folded structures. The folding of ATP was shown also to be enhanced by a three coordination mechanism of Mg2+ with the phosphate group of ATP. This work elucidates the diversity of conformational fluctuations of ADP/Mg:ADP and ATP/Mg:ATP nucleotides and the coordinating dynamics of Mg2+ in molecular motors. oreover, it explores the structural and coordinating changes that take place in F1-ATPase during ATP unbinding.