4.4. Binding Pose MetaDynamics (BPMD)

Binding pose MetaDynamics (BPMD) is an automated, enhanced sampling, metadynamics-based protocol in which the ligand is forced to move around its binding pose. This method showed the ability to reliably discriminate between the correct ligand binding pose and plausible alternatives generated with docking or plain MD studies [18].

According to the protocol, 10 independent metadynamics simulations of 10 ns were performed using as a collective variable (CV) the measure of the root mean square deviation (RMSD) of the ligand heavy atoms with respect to their starting positions. The alignment before the RMSD calculations was done by selecting protein residues within 3 Å of the ligand. The Cαs of these binding site residues were then aligned to those of the first frame of the metadynamics trajectory before calculating the heavy atom RMSD to the ligand conformation in the first frame. The hill height and width were set to 0.05 kcal/ mol (about 1/10 of the characteristic thermal energy of the system, kBT) and 0.02 Å, respectively. Before the actual metadynamics run, the system was solvated in a box of SPC water molecules [38], followed by several minimizations and restrained MD steps that allow the system to slowly reach the desired temperature of 300 K, as well as releasing any bad contacts and/or strain from the initial starting structure. The final snapshot of the short unbiased MD simulation of 0.5 ns was then used as the reference for the following metadynamics production phase. After the simulation, the stability of the ligand during the course was represented by three scores: PoseScore, PersistenceScore (PersScore), and CompositeScore (CompScore). The PoseScore is indicative of the average RMSD from the starting pose. A steep increase of this value is a symptom that the ligand is not in a well-defined energy minimum and, probably, it might not have been accurately modeled. PersScore is a measure of the hydrogen bond persistence calculated in the last 2 ns of the simulation that have the same number of hydrogen bonds as the input structure, averaged over all the 10 repeated simulations. It covers a range between 0 and 1, where 0 indicates that either the starting ligand pose did not have any interactions with the target or that the interactions were lost during the simulations, while 1 indicates that the interactions between the staring ligand pose and the last 2 ns of the simulations were retained. CompositeScore is the linear combination of the PoseScore and PersScore; lower values equate to more stable complexes. Each complex, previously obtained, was run, Country) on a single node with a 1 GPU card NVIDIA GeForce RTX2070.