Fig. 4: Steered molecular dynamics (SMD) or center-of-mass (COM) pulling simulation of C5aR (WT)–PMX53 complex.

a Cut-through section of C5aR (WT)–PMX53–POPC system used for equilibrium MD and steered MD simulations. C5aR is shown in ribbon representation (magenta), embedded in a POPC bilayer (gray) with the headgroup phosphorous atoms shown in sphere representation and the rest of the lipid molecules shown in wire representation. TIP3P water molecules are colored blue, Na⁺ ions purple, and Cl⁻ ions green. Positions and conformations of PMX53 at t = 0 ps, t = 500 ps, and t = 1000 ps derived from the COM pulling simulation are shown in orange, yellow, and dark green colors, respectively. The black arrow is along the z-axis and indicates the direction of pulling of PMX53. b Plot showing force (pN) vs. time (ps) profile obtained for the C5aR (WT)–PMX53 system with a pulling rate of 5 nm ns⁻¹. c Evolution of key intermolecular interactions between C5aR (WT) and PMX53, namely the R6PMX53–D282C5aR salt-bridge (black), and the R6PMX53–Y258C5aR cation-π interaction (red) over the course of the pulling simulation. d Potential of mean force profile calculated for the dissociation of PMX53 from C5aR (WT) using WHAM following umbrella sampling simulations for the C5aR (WT)–PMX53 system. The average PMF profile calculated using bootstrap analysis is presented in the Supplementary Fig. S6f. e Plot showing the number of intermolecular hydrogen bonds (H bonds) formed/broken between the ECL2 region (residues 174–196) of C5aR (WT) and PMX53 over the course of the pulling simulation. f Evolution of key intramolecular interaction R6PMX53–W5PMX53 cation-π interaction (green) in PMX53 over the course of the pulling simulation. g Position and conformation of PMX53 at t = 0 ps during pulling simulation (pull force = 7.62 × 10⁻⁵ pN) where R6PMX53 stably and directly interacts with D282C5aR as compared to the conformation observed in the starting crystal structure conformation. In this conformation, PMX53 forms extensive H bond interactions (shown as black lines) with the residues of C5aR (WT), especially with residues of ECL2. h Position and conformation of PMX53 at t = 325 ps during pulling simulation (pull force = 2386.14 pN) where key non-covalent interactions between PMX53 and C5aR (WT) begin to break and R6PMX53 and W5PMX53 are being pulled away from Y258C5aR and D282C5aR. A number of HBonds between PMX53 and ECL2 also as broken or are in the process of being broken under the influence of the applied force. i Position and conformation of PMX53 at t = 425 ps during pulling simulation (pull force = 879.08 pN) where the PMX53 molecule has been pulled further away with the R6PMX53–D282C5aR salt-bridge and the R6PMX53–Y258C5aR cation-π interaction being completely broken. j Position and conformation of PMX53 at t = 600 ps during pulling simulation (pull force = 29.48 pN) where the ligand is completely unbound from the receptor.