2.4. Molecular Dynamic (MD) Simulations

Molecular Dynamic (MD) simulations provide a robust tool to explore the physical movements of atoms and molecules, thus providing insight into the dynamic evolution of biological systems. The MD simulations were performed using the GPU version of the PMEMD engine provided within the AMBER 18 package [41] ANTECHAMBER was used to generate atomic partial charges for the compounds by utilizing General Amber Force Field (GAFF) procedures. The Leap module of AMBER 18 suspended each system implicitly within an orthorhombic box of TIP3P water molecules ensuring all atoms were within 10Å of any box edge. Leap also allowed for the addition of Na⁺ or Cl^- counter ions for the neutralization of all six systems. An initial minimization of 20000 steps was carried out with an applied restraint potential of 500kcal/mol. This was followed by a full minimization of 10000 steps carried out using the conjugate gradient algorithm in the absence of restraints. A gradual heating MD simulation from 0K to 300K was run for 50ps, to ensure that all systems maintained a fixed number of atoms and volume. The solutes within the system were exposed to a potential harmonic restraint of 10kcal/mol and a collision frequency of 1ps. Following heating, an equilibration step estimating 500ps for each of the systems was conducted with the operating temperature being kept constant at 300K. Additional features such as the number of atoms and pressure were also kept constant, mimicking an isobaric-isothermal ensemble. The systems pressure was maintained at 1 bar using the Berendsen barostat. The total time for the MD simulation conducted was 100ns. In each simulation, the SHAKE algorithm was employed to constrict the bonds of hydrogen atoms. The step size of each simulation was 2fs and an SPFP precision model was used. The simulations coincided with isobaric-isothermal ensemble, with randomized seeding, a constant pressure of 1bar, a pressure-coupling constant of 2ps using the Monte Carlo barostat, a temperature of 300K and Langevin thermostat with collision frequency of 1ps.