Fig 1.
General methodology of our work.
The details are described in the following sections.
Fig 2.
Chemical structure of the main DOR ligands in our study.
(A) Naltrindole (NLT, a δ2-selective morphinan antagonist), (B) DIPP-NH2 (a non-selective peptide δ-antagonist), (C) nalorphine (NLR, δ-non-selective, morphinan partial agonist), (D) KGCHM07 (bifunctional, δ-selective, peptide agonist), (E) DPI287 (δ-selective, benzhydrylpiperazine class agonist), and (F) TIPPψ (δ-selective, pseudopeptide inverse agonist).
Table 1.
General description of our simulated δ systems.
The simulation lengths refer to cMD and GaMD. All the systems were simulated 1 μs as cMD with GROMACS. Then, two replicates of 50 ns as cMD, and finally, 100 ns of GaMD with AMBER18.
Fig 3.
Representative conformers of our seven main simulated DOR systems: Apo-δ, δ-naltrindole, δ-DIPP-NH2, δ-nalorphine, δ-KGCHM07, δ-DPI287, and δ-TIPPψ.
In the apo, naltrindole, DIPP-NH2 and nalorphine complexes, TM5 and TM6 remain distant between them, whilst in the KGCHM07, DPI287, and TIPPψ systems, they are positioned closer to each other at the IC side. H8 experienced unfolding or embedding deeper in the membrane and a large torsion.
Fig 4.
Contact patterns between pairs of TM helices in the seven systems.
(A) The intracellular ends of TM6 and TM7, and (B) the extracellular ends of TM1 and TM2. The active and inactive systems have a lesser number of contacts between TM6 and TM7, while have more contacts between TM1 and TM2.
Fig 5.
Relevant contacts between the ligands in the six complex systems.
(A) The morphinan antagonist naltrindole, (B) the benzhydrylpiperazine agonist DPI287, (C) the pseudopeptide, inverse agonist TIPPψ, and (D) the peptide, full agonist KCGHM07. The uppercase letter A-D identifiers in the snapshots correspond with the rings in the morphinan skeleton. D1283.32 is the anionic counterpart in the mostly conserved -but not exclusive- salt bridge formed with the ligand within the orthosteric site (OSS). Y1293.33 interacts with the phenol function of ring A of the morphinans, tyrosine or dimethyltyrosine (Dmt) moieties; and with the hydrogen-bond-donor in ring C, when the ligand possesses it, as well as K214,5.39 by its hydrophobic and cation parts. H2786.52 also interacts with the phenol group through a water molecule or directly. The transmission switch W2746.48 is in contact with either, the ring A, or the N17-attached group (N-substituent) in the ligand. Q1052.60 interact with the peptide bonds of the peptide bounded agonist ligands, and through hydrophobic contacts with their Phe residues. It is evident that the N-benzyl group of DPI287 tends to interact with N1313.35, and the Tyr1 residue (ring A) in TIPPψ rotates through W2476.48 to a greater extent than the other ligands.
Fig 6.
Morphinan scaffold structures.
(A) Morphinan, (B) 4,5-epoxymorphinan, and (C) morphindole.
Fig 7.
(A) Benzhydrylpiperazine core from DPI287 and BW373U86, and (B) benzhydrylidenepiperidine core from PN6047.
Fig 8.
C-groups attached in the morphinan ligands.
(A) In buprenorphine and other orvinols, (B) in morphindole, and morphobenzofuran ligands, such as naltrindole and SYK657, and (C) benzylidenenaltrexone and SYK656.
Fig 9.
Equivalent scaffold in the peptide ligands, including the pseudopeptide position (ψ) in TIPPψ.
Tic: 1,2,3,4-tetrahydroiso-quinoline-2-carboxylic acid, Dmt: 2,6-dimethyltyrosine.
Fig 10.
Water dynamics in DOR systems.
The cumulative water molecule presence is displayed as blue-colored dots, and the notable regions are highlighted. (A) Apo-δ system, (B) δ-NLT, (C) δ-DIPP-NH2, (D) δ-NLR, (E) δ-KGCHM07, (F) δ-DPI287 y (G) δ-TIPPψ. In the apo and TIPPψ systems, the hydrophobic layer 1 (HL1) is hydrated in the surrounding of H2786.52 and W2746.48, whilst the rest of the systems are dehydrated at this region and under the hydrophobic substructure of the ligands. Only the apo system interacts with a sodium cation at the central coordination site (CCS). At the hydrophobic layer 2 (HL2), the apo, NLT, DIPP-NH2, and NLR systems, there is a wide dehydrated region, whilst in KGCHM07, it is slightly hydrated, and in the DPI287 and TIPPψ complexes, the layer is fully hydrated.
Fig 11.
Coordination of Na+ in the CCS.
A. Minimal distance of any sodium cation to D952.50 and D1283.32 residues, where only the apo-δ system chelates Na+ in the CCS. B. Trajectory between 300 to 500 ns of cMD, showing the path of the Na+ from the EC side to the CCS. C. Representative conformer of the Na+ within CSS, where establishes a hexacoordinated geometry: one coordination bond with S1353.39 and five bonds with water molecules.