Microscopic binding of M5 muscarinic acetylcholine receptor with antagonists by homology modeling, molecular docking, and molecular dynamics simulation

Abstract

By performing homology modeling, molecular docking, and molecular dynamics (MD) simulations, we have developed three-dimensional (3D) structural models of the M5 muscarinic acetylcholine receptor (mAChR) and two complexes for M5 mAChR binding with antagonists SVT-40776 and solifenacin in the environment of lipid bilayer and solvent water. According to the simulated results, each of the antagonists is oriented horizontally in the binding pocket formed by transmembrane helices 2, 3, and 5-7. The cationic headgroup of each of the antagonists interacts with a negatively charged residue, Asp110, through electrostatic and hydrogen-bonding interactions. The simulated results also reveal some significant difference between the binding modes of SVT-40776 and solifenacin. In particular, SVT-40776 is persistently hydrogen bonded with the side chain of residue Tyr458, whereas solifenacin cannot form a similar hydrogen bond with residues around its carbonyl group. Such significant difference in the binding structures is consistent with the fact that SVT-40776 has a much higher binding affinity (K(d) = 0.4 nM) to M5 mAChR than that of solifenacin (K(d) = 31 nM) with the same reeptor. The calculated binding free energy change (-2.3 ± 0.3 kcal/mol) from solifenacin to SVT-40776 is in good agreement with the experimentally derived binding free energy change (-2.58 kcal/mol), suggesting that our modeled M5 mAChR structure and its complexes with the antagonists are reliable. The new structural insights obtained from this computational study are expected to stimulate further biochemical and pharmacological studies on the detailed structures of M5 and other subtypes of mAChRs.

Figure 1

Aligned sequence of M5 mAChR with the template turkey β1-AR. The identified 7 transmembrane (TM) helices and helix 8 are labeled above the sequence. The numbers in parenthesis are the positions for the last residue of TM5. The stars refer to the identical residues, the double filled periods refer to the conservative substitutions, and the filled perigods to the variable conservative substitutions.

Figure 2

Energy-minimized structural model of human M5 mAChR in the physiological environment. (A) View along the normal of the membrane. The M5 mAChR protein is represented as ribbon in rainbow color, lipid molecules in grey, Cl⁻ ions in read spheres, and water molecules in green. (B) Top view of M5 mAChR protein and the lipid molecules. Water molecules are not shown. (C) A hydrophobic packing by residues Ile78, Phe82, and Leu86 from TM2, Trp106 and Leu109 from TM3, and Trp96 from EL-1; (D) An aromatic cluster by residues Phe451, Trp455, and Tyr458 from TM6 and Trp477, Tyr481, and Tyr485 from TM7; (E) Interactions between residues Phe502, Phe506, and Leu509 from H8, Leu50 and Val57 from TM1, and Leu61 from IL-1; (F) Cluster of triple residues. There are three groups of these triple residues, i.e. Leu121-Ser125-Phe374, Leu121-Phe451-Asn491, and Phe451-Asn491-Tyr495. Each of these triple-residues groups could form a Zn²⁺ binding site if they were mutated to residue Histidine and adjust local conformation. The distances among the Cα atoms of each group of triple residues are labeled and represented by lines in different colors. (C) and (D) are in the similar orientation as that of (B), whereas (E) and (F) are the top view from the intracellular side. All residues are colored by the atom type, and the hydrogen atoms are omitted for clarity.

Figure 3

(A) Tracked positional RMSD for SVT-40776 from its original coordinates and tracked distances along the MD trajectory. NH⁺---Asp110(OD1/2) represents the shortest distance between the proton at the cationic head of SVT-40776 and the negatively charged atoms (OD1 or OD2) on the side chain of residue Asp110; O2---Tyr458(HH) is the distance for the hydrogen bond formed by the carbonyl oxygen of SVT-40776 with the Tyr458 side chain; Asp110(OD1/2)---Ser83(HG) represents the distance for the hydrogen bond formed by the Asp110 side chain and the Ser83 side chain; Asp110(OD1/2)---Tyr485(HH) is the distance for the hydrogen bond formed by the Asp110 side chain and the Tyr485 side chain. (B) Top view of representative structure for M5 mAChR-SVT-40776 complex, taken from the last snapshot of the MD simulation. The coloring scheme for the complex structure is the same as that used in Figure 2B, and the SVT-40776 is shown as spheres (left panel) or ball-and-stick (right panel). Residues within 5 Å of SVT-40776 are labeled and shown in stick. The hydrogen bonding interaction between the Asp110 side chain and the cationic head of SVT-40776, the hydrogen bonding interaction between the carbonyl oxygen of SVT-40776 and the Tyr458 side chain, and the hydrogen bonding interactions among side chains of Asp110, Ser83, and Tyr485 are shown in dashed lines along with the labeled distances.

Figure 4

(A) Plots of the tracked positional RMSD for Solifenacin from its original coordinates and tracked distances. NH⁺---Asp110(OD1/2) represents the shortest distance between the proton at the cationic head of Solifenacin and the negatively charged atoms (OD1 or OD2) on the side chain of residue Asp110; Asp110(OD1/2)---Ser83(HG) represents the distance for the hydrogen bond formed by the Asp110 side chain and the Ser83 side chain; Asp110(OD1/2)---Tyr485(HH) is the distance for the hydrogen bond formed by the Asp110 side chain and the Tyr485 side chain. (B) Top view of representative structure for M5 mAChR-Solifenacin complex, taken from the last snapshot of the MD simulations. The coloring scheme for the complex structure is the same as that used in Figure 2B, and the Solifenacin is presented as spheres (left panel) or ball-and-stick (right panel). Residues within 5 Å of Solifenacin are labeled and shown in stick. The hydrogen bonding interaction between the Asp110 side chain and the cationic head of Solifenacin, and the hydrogen bonding interactions among side chains of Asp110, Ser83, and Tyr485 are shown in dashed lines along with the labeled distances.

Scheme 1