Figure 1.
Structure of Amarogentin, a secoiridoid glycoside from Swertia chirayita.
Amarogentin consists of three essential subgroups, the iridoid group, the glucose moiety and the biphenyl-triol rings.
Table 1.
AutoDock binding energy values and H-bond forming residues of the lead molecules.
Figure 2.
RMSD of the backbone of COX-1 and COX-2 modelled proteins after stabilisation.
Protein backbone RMSD of COX-1 model, over a time frame of 15ns (in black), shows stability in the last 5ns time frame, deviating about 0.35nm from the native structure whereas, protein backbone RMSD of COX-2 model, for 10ns (in red), achieved stability after 5ns and maintained till the final 10ns, with an average deviation of about 0.3nm. The plot has been generated using the GRACE plotting tool.
Figure 3.
Protein stability assessment in terms of Rg and RMSF.
(A) The overall movement in the protein, with respect to the initial frame, is calculated using Rg plot. Rg plot for COX-1 (in black) showed an average deviation of 2.34nm and an average deviation of 2.30nm for COX-2 (in red). (B) Residue fluctuations were observed using the RMSF plot and the overall fluctuation was found to be marginal for both the proteins, with a maximum deviation occurring along the terminal residues and the loop region. The active site residues showed intactness for both COX-1 (black) and COX-2 (red) protein models with very low observed fluctuations. The plots have been generated using the GRACE plotting tool.
Figure 4.
2D representation of the docking pose of amarogentin inside the hydrophobic cavity of COX.
(A) The docking pose of amarogentin inside the binding cavity of COX-1 shows that besides forming several van der Waals interactions, amarogentin forms four H-bonds, two with Arg119 and one with Ser352. (B) A total of five H-bonds were also observed in the docked amarogentin-COX-2 complex, one each with Arg106, Ser516, Tyr371, and Met508. Amarogentin formed van der Waals interaction with a several residues along the hydrophobic channel. (Legend: green spheres represent hydrophobic residues; cyan spheres represent residues with polar side chains; red spheres and purple spheres show negatively and positively charged amino acid residues, respectively; solid red lines denote cation- π interaction; dotted purple lines indicate hydrogen bonds with side-chain atoms, with the direction of arrow denoting the acceptor atom; solid green lines indicate π-π interactions; and the grey spheres surrounding the atoms indicate that the atoms are exposed to solvent). The illustrations have been generated using Schrödinger Maestro open-source visualisation package.
Figure 5.
RMSD of amarogentin-COX complexes for 40ns time frame.
RMSD of amarogentin-COX-1 complex (black) shows an elevation in the deviation from the initial structure, reaching around 0.3nm for the final 40ns frame. The RMSD of amarogentin-COX-2 complex (red) showed initial deviations but attained stability at 15ns and remained so till the final 40ns time frame with an RMSD of 0.18nm. The plots have been generated using the GRACE plotting tool.
Figure 6.
Interaction of amarogentin with COX-1 based on its position at 0ns and 40ns.
(A) Interaction of amarogentin with channel gate forming residues, at 0ns (red) and 40ns (yellow). The movement in the position and angle of the residue, at 0ns (green) and 40ns (cyan), can differentiate their deviations and suggest a movement of amarogentin outside the channel breaching the channel gate. (B) Position of amarogentin at 0ns (red) and at 40ns (yellow) clearly indicates a complete shift in its orientation, in the course of the simulation. The figure was generated using the PyMol molecular visualisation tool.
Figure 7.
2D representation of the binding pose of amarogentin inside the hydrophobic cavity of COX isoforms after 40ns simulation.
(A) Amarogentin formed only one H-bond with Ser352 in the COX-1 hydrophobic cavity. However, atoms which formed H-bonds after docking were present as van der Waal contacts. (B) Amarogentin made a total of six hydrogen bonds and was positioned up in the channel making the pose look more stable after simulation. Legends are same as expressed in Figure 4. The illustrations have been generated using Schrödinger Maestro open-source visualisation package.
Figure 8.
Interaction of amarogentin with COX-2 based on its position at 0ns and 40ns.
(A) Interaction of amarogentin with channel gate forming residues, at 0ns (red) and 40ns (yellow). There is comparatively less deviation in the side chains of channel gate residues at 0ns (green) and at 40ns (cyan). (B) Change in the position of amarogentin at 0ns (red) and at 40ns (yellow) indicate a marginal deviation in its structure at 40ns from the native 0ns structure. The illustrations were generated using the PyMol molecular visualisation tool.
Table 2.
Comparison of calculated Binding free energies for COX-2 selective drugs with known experimental data.
Figure 9.
Movement of amarogentin inside the COX-1 binding cavity.
Amarogentin after 40ns (green) simulation shows a clear movement outside the binding channel (grey) with respect to its initial 0ns frame (purple), indicating that the complex may not be stable in nature. The figure was generated using the PyMol molecular visualisation tool.
Table 3.
Overall Non-bonded Interaction profile of amarogentin with COX-1 and COX-2.