Class C G protein-coupled receptors (GPCRs) regulate important physiological functions and allosteric modulators binding to the transmembrane domain constitute an attractive and, due to a lack of structural insight, a virtually unexplored potential for therapeutics and the food industry. Combining pharmacological site-directed mutagenesis data with the recent class C GPCR experimental structures will provide a foundation for rational design of new therapeutics.We uncover one common site for both positive and negative modulators with different amino acid layouts that can be utilized to obtain selectivity. Additionally, we show a large potential for structure-based modulator design, especially for four orphan receptors with high similarity to the crystal structures.All collated mutagenesis data is available in the GPCRdb mutation browser at http://gpcrdb.org/mutations/ and can be analyzed online or downloaded in excel format.david.gloriam@sund.ku.dk.Supplementary data are available at Bioinformatics online.
Serotonergic psychedelics are described to have activation of the serotonin 2A receptor (5-HT2A) as their main pharmacological action. Despite their relevance, the molecular mechanisms underlying the psychedelic effects induced by certain 5-HT2A agonists remain elusive. One of the proposed hypotheses is the occurrence of biased agonism, defined as the preferential activation of certain signaling pathways over others. This study comparatively monitored the efficiency of a diverse panel of 4-position-substituted (and N-benzyl-derived) phenylalkylamines to induce recruitment of β-arrestin2 (βarr2) or miniGαq to the 5-HT2A, allowing us to assess structure–activity relationships and biased agonism. All test compounds exhibited agonist properties with a relatively large range of both EC50 and Emax values. Interestingly, the lipophilicity of the 2C-X phenethylamines was correlated with their efficacy in both assays but yielded a stronger correlation in the miniGαq- than in the βarr2-assay. Molecular docking suggested that accommodation of the 4-substituent of the 2C-X analogues in a hydrophobic pocket between transmembrane helices 4 and 5 of 5-HT2A may contribute to this differential effect. Aside from previously used standard conditions (lysergic acid diethylamide (LSD) as a reference agonist and a 2 h activation profile to assess a compound's activity), serotonin was included as a second reference agonist, and the compounds' activities were also assessed using the first 30 min of the activation profile. Under all assessed circumstances, the qualitative structure–activity relationships remained unchanged. Furthermore, the use of two reference agonists allowed for the estimation of both "benchmark bias" (relative to LSD) and "physiology bias" (relative to serotonin).
The N-benzylphenethylamines (NBOMes) are a class of ligands from which compounds with impressive selectivity for the serotonin 2A receptor (5-HT2AR) over the closely related serotonin 2C receptor (5-HT2CR) have emerged. These include 4-(2-((2-hydroxybenzyl)amino)ethyl)-2,5-dimethoxybenzonitrile (25CN-NBOH, 1) and 2-(2,5-dimethoxy-4-bromobenzyl)-6-(2-methoxyphenyl)piperidine (DMPMBB, 2). The present work entails the synthesis and characterization of ligands wherein the structures of these two molecules have been fused. The desired compounds were accessed by a six-step synthetic procedure followed by the chiral resolution of the resulting racemic mixtures, giving one active ((S,S)-3) and three essentially inactive stereoisomers. In silico experiments support that one of the four possible stereoisomers would be active. Further in silico investigations showed that 1, 2, and (S,S)-3 share a common binding mode, further supporting the shared stereochemistry between the active enantiomer ((S,S)-3) and 2.
γ-Hydroxybutyric acid (GHB) is a neuroactive substance with specific high-affinity binding sites. To facilitate target identification and ligand optimization, we herein report a comprehensive structure-affinity relationship study for novel ligands targeting these binding sites. A molecular hybridization strategy was used based on the conformationally restricted 3-hydroxycyclopent-1-enecarboxylic acid (HOCPCA) and the linear GHB analog trans-4-hydroxycrotonic acid (T-HCA). In general, all structural modifications performed on HOCPCA led to reduced affinity. In contrast, introduction of diaromatic substituents into the 4-position of T-HCA led to high-affinity analogs (medium nanomolar Ki) for the GHB high-affinity binding sites as the most high-affinity analogs reported to date. The SAR data formed the basis for a three-dimensional pharmacophore model for GHB ligands, which identified molecular features important for high-affinity binding, with high predictive validity. These findings will be valuable in the further processes of both target characterization and ligand identification for the high-affinity GHB binding sites.
The binding affinity of a drug-like molecule depends among other things on the availability of the bioactive conformation. If the bioactive conformation has a significantly higher energy than the global minimum energy conformation, then the molecule is unlikely to bind to its target. Determination of the global minimum energy conformation and calculation of conformational penalties of binding is a prerequisite for prediction of reliable binding affinities. Here, we present a simple and computationally efficient procedure to estimate the global energy minimum for a wide variety of structurally diverse molecules, including polar and charged compounds. Identifying global energy minimum conformations of such compounds with force field methods is problematic due to the exaggeration of intramolecular electrostatic interactions. We demonstrate that the global energy minimum conformations of zwitterionic compounds generated by conformational analysis with modified electrostatics are good approximations of the conformational distributions predicted by experimental data and with molecular dynamics performed in explicit solvent. Finally the method is used to calculate conformational penalties for zwitterionic GluA2 agonists and to filter false positives from a docking study.
Peptides targeting disease-relevant protein–protein interactions are an attractive class of therapeutics covering the otherwise undruggable space between small molecules and therapeutic proteins. However, peptides generally suffer from poor metabolic stability and low membrane permeability. Hence, peptide cyclization has become a valuable approach to develop linear peptide motifs into metabolically stable and potentially cell-permeable cyclic leads. Furthermore, cyclization of side chains, also known as "stapling", can stabilize particular secondary peptide structures. Here, we demonstrate that a comprehensive examination of cyclization strategies in terms of position, chemistry, and length is a prerequisite for the selection of optimal cyclic peptide scaffolds. Our systematic approach identifies cyclic APP dodecamer peptides targeting the phosphotyrosine binding domain of Mint2 with substantially improved affinity. We show that especially all-hydrocarbon stapling provides improved metabolic stability, a significantly stabilized secondary structure and membrane permeability.
'/%3e%3cuse xlink:href='%23a' transform='rotate(23.036 .093 25.536)'/%3e%3cuse xlink:href='%23a' transform='rotate(45.87 1.273 16.18)'/%3e%3cuse xlink:href='%23a' transform='rotate(69.945 .996 12.078)'/%3e%3cuse xlink:href='%23a' transform='rotate(20.66 -19.689 31.932)'/%3e%3c/svg%3e)
'/%3e%3c/defs%3e%3cpath fill='%23012169' d='M0 0h10080v5040H0z'/%3e%3cpath stroke='%23fff' stroke-width='.6' d='m0 0 6 3m0-3L0 3' clip-path='url(%23b)' transform='scale(840)'/%3e%3cpath stroke='%23e4002b' stroke-width='.4' d='m0 0 6 3m0-3L0 3' clip-path='url(%23c)' transform='scale(840)'/%3e%3cpath stroke='%23fff' stroke-width='840' d='M2520 0v2520M0 1260h5040'/%3e%3cpath stroke='%23e4002b' stroke-width='504' d='M2520 0v2520M0 1260h5040'/%3e%3cg fill='%23fff'%3e%3cuse xlink:href='%23d' x='2520' y='3780'/%3e%3cuse xlink:href='%23a' x='7560' y='4200'/%3e%3cuse xlink:href='%23a' x='6300' y='2205'/%3e%3cuse xlink:href='%23a' x='7560' y='840'/%3e%3cuse xlink:href='%23a' x='8680' y='1869'/%3e%3cuse xlink:href='%23e' x='8064' y='2730'/%3e%3c/g%3e%3c/svg%3e)