Certain types of nonpsychoactive cannabinoids can potentiate glycine receptors (GlyRs), an important target for nociceptive regulation at the spinal level. However, little is known about the ...potential and mechanism of glycinergic cannabinoids for chronic pain treatment. We report that systemic and intrathecal administration of cannabidiol (CBD), a major nonpsychoactive component of marijuana, and its modified derivatives significantly suppress chronic inflammatory and neuropathic pain without causing apparent analgesic tolerance in rodents. The cannabinoids significantly potentiate glycine currents in dorsal horn neurons in rat spinal cord slices. The analgesic potency of 11 structurally similar cannabinoids is positively correlated with cannabinoid potentiation of the α3 GlyRs. In contrast, the cannabinoid analgesia is neither correlated with their binding affinity for CB1 and CB2 receptors nor with their psychoactive side effects. NMR analysis reveals a direct interaction between CBD and S296 in the third transmembrane domain of purified α3 GlyR. The cannabinoid-induced analgesic effect is absent in mice lacking the α3 GlyRs. Our findings suggest that the α3 GlyRs mediate glycinergic cannabinoid-induced suppression of chronic pain. These cannabinoids may represent a novel class of therapeutic agents for the treatment of chronic pain and other diseases involving GlyR dysfunction.
The neuronal α4β2 nicotinic acetylcholine receptor (nAChR) is a target for general anesthetics. Currently available experimental structural information is inadequate to understand where anesthetics ...bind and how they modulate the receptor motions essential to function. Using our published open-channel structure model of α4β2 nAChR, we identified and evaluated six amphiphilic interaction sites for the volatile anesthetic halothane via flexible ligand docking and subsequent 20-ns molecular dynamics simulations. Halothane binding energies ranged from −6.8 to −2.4 kcal/mol. The primary binding sites were located at the interface of extracellular and transmembrane domains, where halothane perturbed conformations of, and widened the gap among, the Cys loop, the β1−β2 loop, and the TM2−TM3 linker. The halothane with the highest binding affinity at the interface between the α4 and β2 subunits altered interactions between the protein and nearby lipids by competing for hydrogen bonds. Gaussian network model analyses of the α4β2 nAChR structures at the end of 20-ns simulations in the absence or presence of halothane revealed profound changes in protein residue mobility. The concerted motions critical to protein function were also perturbed considerably. Halothane’s effect on protein dynamics was not confined to the residues adjacent to the binding sites, indicating an action on a more global scale.
Systemic and intrathecal administration of derivatives of a nonpsychoactive component of marijuana significantly suppresses chronic inflammatory and neuropathic pain, without causing analgesic ...tolerance, in several rodent models.
Three mechanistic possibilities for the formation of the oxetane (D ring) of Taxol were examined at various levels of theory B3LYP/6-31+G(d,p), mPW1PW91/6-31+G(d,p), and MP2/6-31+G(d,p) including one ...mechanism involving an unusual oxabicyclobutonium ion intermediate. The mechanisms examined differ considerably in terms of their predicted inherent activation barriers, and the requirements for acceleration of each by an enzyme active site are outlined. Our calculations provide an important starting point for future studies in this area. Also examined were previously published calculations on simple oxabicyclobutonium ions, as well as the all-carbon analog of the pathway involving the oxabicyclobutonium ion.
The anesthetic propofol inhibits the currents of the homopentameric ligand-gated ion channel GLIC, yet the crystal structure of GLIC with five propofol molecules bound symmetrically shows an ...open-channel conformation. To address this dilemma and determine if the symmetry of propofol binding sites affects the channel conformational transition, we performed a total of 1.5 μs of molecular dynamics simulations for different GLIC systems with propofol occupancies of 0, 1, 2, 3, and 5. GLIC without propofol binding or with five propofol molecules bound symmetrically, showed similar channel conformation and hydration status over multiple replicates of 100-ns simulations. In contrast, asymmetric binding to one, two or three equivalent sites in different subunits accelerated the channel dehydration, increased the conformational heterogeneity of the pore-lining TM2 helices, and shifted the lateral and radial tilting angles of TM2 toward a closed-channel conformation. The results differentiate two groups of systems based on the propofol binding symmetry. The difference between symmetric and asymmetric groups is correlated with the variance in the propofol-binding cavity adjacent to the hydrophobic gate and the force imposed by the bound propofol. Asymmetrically bound propofol produced greater variance in the cavity size that could further elevate the conformation heterogeneity. The force trajectory generated by propofol in each subunit over the course of a simulation exhibits an ellipsoidal shape, which has the larger component tangential to the pore. Asymmetric propofol binding creates an unbalanced force that expedites the channel conformation transitions. The findings from this study not only suggest that asymmetric binding underlies the propofol functional inhibition of GLIC, but also advocate for the role of symmetry breaking in facilitating channel conformational transitions.
Pentameric ligand-gated ion channels (pLGICs) are targets of general anesthetics, but a structural understanding of anesthetic action on pLGICs remains elusive. GLIC, a prokaryotic pLGIC, can be ...inhibited by anesthetics, including ketamine. The ketamine concentration leading to half-maximal inhibition of GLIC (58 μM) is comparable to that on neuronal nicotinic acetylcholine receptors. A 2.99 Å resolution X-ray structure of GLIC bound with ketamine revealed ketamine binding to an intersubunit cavity that partially overlaps with the homologous antagonist-binding site in pLGICs. The functional relevance of the identified ketamine site was highlighted by profound changes in GLIC activation upon cysteine substitution of the cavity-lining residue N152. The relevance is also evidenced by changes in ketamine inhibition upon the subsequent chemical labeling of N152C. The results provide structural insight into the molecular recognition of ketamine and are valuable for understanding the actions of anesthetics and other allosteric modulators on pLGICs.
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► The structure of ketamine-bound GLIC reveals an anesthetic binding site ► The study provides compelling evidence for allosteric inhibition by anesthetics ► Ketamine inhibition on GLIC is similar to competitive antagonist action on nAChRs ► Ketamine directly acts on pLGICs in addition to NMDA receptors
Pentameric ligand-gated ion channels (pLGICs) are targets of general anesthetics (GAs), but a structural basis of GA action remains elusive. Pan et al. describe a structure of a prokaryotic pLGIC in complex with GA, ketamine, which shows GA binding to a cavity different from the orthosteric agonist binding site.
Pentameric ligand-gated ion channels are targets of general anesthetics. Although the search for discrete anesthetic binding sites has achieved some degree of success, little is known regarding how ...anesthetics work after the events of binding. Using the crystal structures of the bacterial Gloeobacter violaceus pentameric ligand-gated ion channel (GLIC), which is sensitive to a variety of general anesthetics, we performed multiple molecular dynamics simulations in the presence and absence of the general anesthetic isoflurane. Isoflurane bound to several locations within GLIC, including the transmembrane pocket identified crystallographically, the extracellular (EC) domain, and the interface of the EC and transmembrane domains. Isoflurane also entered the channel after the pore was dehydrated in one of the simulations. Isoflurane disrupted the quaternary structure of GLIC, as evidenced in a striking association between the binding and breakage of intersubunit salt bridges in the EC domain. The pore-lining helix experienced lateral and inward radial tilting motion that contributed to the channel closure. Isoflurane binding introduced strong anticorrelated motions between different subunits of GLIC. The demonstrated structural and dynamical modulations by isoflurane aid in the understanding of the underlying mechanism of anesthetic inhibition of GLIC and possibly other homologous pentameric ligand-gated ion channels.
Quantum chemical calculations are used to assess various means of lowering the barrier for the dyotropic rearrangement previously proposed to occur during the carbocation rearrangement process ...promoted by pentalenene synthase. Several means of lowering this barrier, including a stepwise pathway for dyotropic rearrangement, are uncovered.
The neuronal α4β2 nicotinic acetylcholine receptor (nAChR) is a potential molecular target for general anesthetics. It is unclear, however, whether anesthetic action produces the same effect on the ...open and closed channels. Computations parallel to our previous open channel study (J. Phys. Chem. B 2009, 113, 12581) were performed on the closed-channel α4β2 nAChR to investigate the conformation-dependent anesthetic effects on channel structures and dynamics. Flexible ligand docking and over 20 ns molecular dynamics simulations revealed similar halothane-binding sites in the closed and open channels. The sites with relatively high binding affinities (∼ −6.0 kcal/mol) were identified at the interface of extracellular (EC) and transmembrane (TM) domains or at the interface between α4 and β2 subunits. Despite similar sites for halothane binding, the closed-channel conformation showed much less sensitivity than the open channel to the structural and dynamical perturbations from halothane. Compared to the systems without anesthetics, the amount of water inside the pore decreased by 22% in the presence of halothane in the open channel but only by 6% in the closed channel. Comparison of the nonbonded interactions at the EC/TM interfaces suggested that the β2 subunits were more prone than the α4 subunits to halothane binding. In addition, our data support the notion that halothane exerts its effect by disturbing the quaternary structure and dynamics of the channel. The study concludes that sensitivity and global dynamics responsiveness of α4β2 nAChR to halothane are conformation dependent. The effect of halothane on the global dynamics of the open-channel conformation might also account for the action of other inhaled general anesthetics.
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