Then the seven complexes GLOI indomethacin

Then the seven complexes (GLOI-indomethacin, GLOI-zomepirac, GLOI-fenoprofen, and GLOI-ketoprofen, GLOI-tolmetin GLOI-curcumin, and GLOI-bisdemethoxycurcumin) were energy-minimized to remove unfavorable steric strain relative to the force field adopted. Further MD simulations were carried out based on the relaxed structures. Each complex was progressively heated from 0 to 300K in 50ps and equilibrated for another 50ps at 300K using the NVT (constant composition, volume, and temperature) ensemble with a weak constraint of 10kcalmolÅ. Finally, periodic boundary dynamics simulations of 6ns were carried out for the production step in an NPT (constant composition, pressure, and temperature) ensemble at 1atm and 300K. The temperature was controlled by the weak-coupling algorithm, while the long-range electrostatic interactions were treated by the Particle-Mesh-Ewald method with a non-bonded cut-off of 10Å. All the simulations proceeded with the SHAKE algorithm turned on. Output trajectory files were sampled every 2ps for subsequent analysis. The MM-PBSA approach, , , , , , encoded in AMBER was used to estimate the binding free everolimus (Δ) in continuum solvent representation of the protein-ligand systems. A total of 100 snapshots were taken from each last 1ns trajectory with an interval of 10ps. The entropy contribution was not considered because the structural similarity of the inhibitors made insignificant differences at the studied temperature. The detailed information about the MM-PBSA approach is provided in . The kinetic inhibition constants () of five NASIDs towards GLOI have been determined by others previously. The inhibitory affinities (see the curves in ) of four NASIDs except tolmetin were reassessed under identical conditions in the present work. As shown in , , our experimental results lead to a consistent conclusion with those in the literature. To validate the dynamic stability of the model systems, the RMSD values for the backbone atoms of the 6-ns MD trajectories were monitored using the X-ray crystal structure as a reference. shows that the RMSD values of the seven complexes reached convergence within 2Å, which ensure the dynamic stability of MD trajectories of the complexes. Therefore, the subsequent analyses were based on the MD trajectories between 5 and 6ns. . The predicted binding free energies Δ (including the individual energy term) were calculated by the MM-PBSA method (). Based on Δ and the approximately estimated Δ from our experimental values (via ≈ RT ln ), our statistical analysis achieved a significantly linear correlation () with a high conventional regression coefficient (=0.974). This suggests that Δ of a GLOI/inhibitor complex can be used to prognosticate the experimental inhibitory affinity of a ligand towards GLOI of similar structural features. . Brown et al. developed a series of plots to serve as a reality check for models of a certain number of data points and potency span. For the dataset of this model, which spans a potency range of 2log units and has six points, it is estimated that the correlation coefficient is likely to be between 0.8 and 0.9 in terms of experimental error only (=0.3) according to the research. Thus, our correlation coefficient of 0.974 is over expectation. However according to the statistic analyses in the same work, among 16 publications from 2006 to 2007, 12 of the values of them were better than expected and two of them were worse than expected. Only two are in the expected range. This result exemplified that it is a usual case in which values can be out of the reasonable range. . Several molecular properties such as molecular weight, log, p, were used to study the dependence of actual binding affinities on molecular properties. However, none of them produced better correlations () than that of the predicted binding energy by the MM-PBSA method since those molecular properties do not take the receptor GLOI into consideration. Δ Each of the energy terms involved in Δ represents a specific contribution to the total binding free energy. Among the four terms (Δ, Δ, Δ, and Δ), the internal electrostatic one Δ yields a reasonable correlation () with which emphasizes the relative importance of the internal electrostatic interactions in the inhibition of these ligands against GLOI. The studied GLOI inhibitors could be broadly divided into two groups in terms of Δ. Group-A is represented by indomethacin, tolmetin, curcumin, and bisdemethoxycurcumin, while Group-B by zomepirac, fenoprofen, and ketoprofen. The internal electrostatic energies of Group-A are remarkably more dominating than those of Group-B. These energy differences of these two groups reflect diverse binding modes of these ligands.