RÉSUMÉ
Adenosine A2A receptors (A2AAR) evoke pleiotropic intracellular signaling events via activation of the stimulatory heterotrimeric G protein, Gs. Here, we used cryoEM to solve the agonist-bound structure of A2AAR in a complex with full-length Gs α and Gß4γ2 (A2AAR-Gs α:ß4γ2). The orthosteric binding site of A2AAR-Gs α:ß4γ2 was similar to other structures of agonist-bound A2AAR, with or without Gs. Unexpectedly, the solvent accessible surface area within the interior of the complex was substantially larger for the complex with Gß4 versus the closest analog, A2AAR-miniGs α:ß1γ2. Consequently, there are fewer interactions between the switch II in Gs α and the Gß4 torus. In reconstitution experiments Gß4γ2 displayed a ten-fold higher efficiency over Gß1γ2 in catalyzing A2AAR dependent GTPγS binding to Gs α. We propose that the less constrained switch II in A2AAR-Gs α:ß4γ2 accounts for this increased efficiency. These results suggest that Gß4 functions as a positive allosteric enhancer versus Gß1.
RÉSUMÉ
The technique of cryogenic-electron microscopy (cryo-EM) has revolutionized the field of membrane protein structure and function with a focus on the dominantly observed molecular species. This report describes the structural characterization of a fully active human apelin receptor (APJR) complexed with heterotrimeric G protein observed in both 2:1 and 1:1 stoichiometric ratios. We use cryo-EM single-particle analysis to determine the structural details of both species from the same sample preparation. Protein preparations, in the presence of the endogenous peptide ligand ELA or a synthetic small molecule, both demonstrate these mixed stoichiometric states. Structural differences in G protein engagement between dimeric and monomeric APJR suggest a role for the stoichiometry of G protein-coupled receptor- (GPCR-)G protein coupling on downstream signaling and receptor pharmacology. Furthermore, a small, hydrophobic dimer interface provides a starting framework for additional class A GPCR dimerization studies. Together, these findings uncover a mechanism of versatile regulation through oligomerization by which GPCRs can modulate their signaling.
Sujet(s)
Protéines G , Récepteurs couplés aux protéines G , Récepteur de l'apeline/composition chimique , Récepteur de l'apeline/métabolisme , Protéines de transport/métabolisme , Protéines G/métabolisme , Humains , Récepteurs couplés aux protéines G/composition chimique , Transduction du signalRÉSUMÉ
Glucagon-like peptide-1 receptor (GLP-1R) is a class B G protein-coupled receptor that plays an important role in glucose homeostasis and treatment of type 2 diabetes. Structures of full-length class B receptors were determined in complex with their orthosteric agonist peptides, however, little is known about their extracellular domain (ECD) conformations in the absence of orthosteric ligands, which has limited our understanding of their activation mechanism. Here, we report the 3.2 Å resolution, peptide-free crystal structure of the full-length human GLP-1R in an inactive state, which reveals a unique closed conformation of the ECD. Disulfide cross-linking validates the physiological relevance of the closed conformation, while electron microscopy (EM) and molecular dynamic (MD) simulations suggest a large degree of conformational dynamics of ECD that is necessary for binding GLP-1. Our inactive structure represents a snapshot of the peptide-free GLP-1R and provides insights into the activation pathway of this receptor family.
Sujet(s)
Récepteur du peptide-1 similaire au glucagon/composition chimique , Séquence d'acides aminés , Apoprotéines/composition chimique , Disulfures/métabolisme , Récepteur du peptide-1 similaire au glucagon/ultrastructure , Humains , Ligands , Simulation de dynamique moléculaire , Conformation des protéines , Stabilité protéique , Récepteurs au glucagon/composition chimiqueRÉSUMÉ
Selective activation of the δ-opioid receptor (DOP) has great potential for the treatment of chronic pain, benefitting from ancillary anxiolytic and antidepressant-like effects. Moreover, DOP agonists show reduced adverse effects as compared to µ-opioid receptor (MOP) agonists that are in the spotlight of the current "opioid crisis." Here, we report the first crystal structures of the DOP in an activated state, in complex with two relevant and structurally diverse agonists: the potent opioid agonist peptide KGCHM07 and the small-molecule agonist DPI-287 at 2.8 and 3.3 Å resolution, respectively. Our study identifies key determinants for agonist recognition, receptor activation, and DOP selectivity, revealing crucial differences between both agonist scaffolds. Our findings provide the first investigation into atomic-scale agonist binding at the DOP, supported by site-directed mutagenesis and pharmacological characterization. These structures will underpin the future structure-based development of DOP agonists for an improved pain treatment with fewer adverse effects.
Sujet(s)
Simulation de docking moléculaire , Peptides/composition chimique , Récepteur delta/agonistes , Récepteur delta/composition chimique , Animaux , Cristallographie aux rayons X , Humains , Domaines protéiques , Récepteur mu/agonistes , Récepteur mu/composition chimique , Cellules Sf9 , SpodopteraRÉSUMÉ
G-protein-coupled receptors (GPCRs) transduce physiological and sensory stimuli into appropriate cellular responses and mediate the actions of one-third of drugs. GPCR structural studies have revealed the general bases of receptor activation, signaling, drug action and allosteric modulation, but so far cover only 13% of nonolfactory receptors. We broadly surveyed the receptor modifications/engineering and methods used to produce all available GPCR crystal and cryo-electron microscopy (cryo-EM) structures, and present an interactive resource integrated in GPCRdb ( http://www.gpcrdb.org ) to assist users in designing constructs and browsing appropriate experimental conditions for structure studies.
Sujet(s)
Biologie informatique/méthodes , Internet , Récepteurs couplés aux protéines G/génétique , Site allostérique , Animaux , Bovins , Cryomicroscopie électronique , Cristallographie aux rayons X , Bases de données de protéines , Conception de médicament , Glycosylation , Cellules HEK293 , Humains , Mutation , Phosphorylation , Domaines protéiques , Ingénierie des protéines , Rhodopsine/composition chimique , Transduction du signal , LogicielRÉSUMÉ
Misoprostol is a life-saving drug in many developing countries for women at risk of post-partum hemorrhaging owing to its affordability, stability, ease of administration and clinical efficacy. However, misoprostol lacks receptor and tissue selectivities, and thus its use is accompanied by a number of serious side effects. The development of pharmacological agents combining the advantages of misoprostol with improved selectivity is hindered by the absence of atomic details of misoprostol action in labor induction. Here, we present the 2.5 Å resolution crystal structure of misoprostol free-acid form bound to the myometrium labor-inducing prostaglandin E2 receptor 3 (EP3). The active state structure reveals a completely enclosed binding pocket containing a structured water molecule that coordinates misoprostol's ring structure. Modeling of selective agonists in the EP3 structure reveals rationales for selectivity. These findings will provide the basis for the next generation of uterotonic drugs that will be suitable for administration in low resource settings.
Sujet(s)
Misoprostol/composition chimique , Sous-type EP3 des récepteurs des prostaglandines E/composition chimique , Sous-type EP3 des récepteurs des prostaglandines E/métabolisme , Sites de fixation , Cristallographie aux rayons X , Dinoprostone/analogues et dérivés , Dinoprostone/composition chimique , Dinoprostone/métabolisme , Humains , Misoprostol/métabolisme , Simulation de docking moléculaire , Mutagenèse dirigée , Conformation des protéines , Sous-type EP3 des récepteurs des prostaglandines E/agonistes , Sous-type EP3 des récepteurs des prostaglandines E/génétique , Transduction du signal , Eau/composition chimiqueRÉSUMÉ
In the version of this article originally published, the present address for Petr Popov was incorrectly listed as 'Koltech Institute of Science & Technology, Moscow, Russia'. The correct present address is 'Skolkovo Institute of Science and Technology, Moscow, Russia'. The error has been corrected in the HTML and PDF versions of the paper.
RÉSUMÉ
G protein-coupled receptors (GPCRs) represent the largest class of cell surface proteins and thus constitute an important family of therapeutic targets. Therefore, significant effort has been put towards the identification of novel ligands that can modulate the activity of a GPCR target with high efficacy and selectivity. However, due to limitations inherent to the most common techniques for GPCR ligand discovery, there is a pressing need for more efficient and effective ligand screening methods especially for the identification of potential allosteric modulators. Here we present a high-throughput, label-free and unbiased screening approach for the identification of small molecule ligands towards GPCR targets based on affinity mass spectrometry. This new approach features the usage of target-expressing cell membranes rather than purified proteins for ligand screening and allows the detection of both orthosteric and allosteric ligands targeting specific GPCRs. Screening a small compound library with this approach led to the rapid discovery of an antagonist for the 5-HT receptor and four positive allosteric modulators for GLP-1 receptor that were not previously reported.
RÉSUMÉ
Drugs frequently require interactions with multiple targets-via a process known as polypharmacology-to achieve their therapeutic actions. Currently, drugs targeting several serotonin receptors, including the 5-HT2C receptor, are useful for treating obesity, drug abuse, and schizophrenia. The competing challenges of developing selective 5-HT2C receptor ligands or creating drugs with a defined polypharmacological profile, especially aimed at G protein-coupled receptors (GPCRs), remain extremely difficult. Here, we solved two structures of the 5-HT2C receptor in complex with the highly promiscuous agonist ergotamine and the 5-HT2A-C receptor-selective inverse agonist ritanserin at resolutions of 3.0 Å and 2.7 Å, respectively. We analyzed their respective binding poses to provide mechanistic insights into their receptor recognition and opposing pharmacological actions. This study investigates the structural basis of polypharmacology at canonical GPCRs and illustrates how understanding characteristic patterns of ligand-receptor interaction and activation may ultimately facilitate drug design at multiple GPCRs.
Sujet(s)
Ergotamine/composition chimique , Récepteur de la sérotonine de type 5-HT2C/composition chimique , Ritansérine/composition chimique , Agonistes des récepteurs 5-HT2 de la sérotonine/composition chimique , Antagonistes des récepteurs 5-HT2 de la sérotonine/composition chimique , Cellules HEK293 , Humains , Obésité/traitement médicamenteux , Obésité/métabolisme , Domaines protéiques , Récepteur de la sérotonine de type 5-HT2C/métabolisme , Schizophrénie/traitement médicamenteux , Schizophrénie/métabolisme , Relation structure-activité , Troubles liés à une substance/traitement médicamenteux , Troubles liés à une substance/métabolismeRÉSUMÉ
Class B G-protein-coupled receptors (GPCRs), which consist of an extracellular domain (ECD) and a transmembrane domain (TMD), respond to secretin peptides to play a key part in hormonal homeostasis, and are important therapeutic targets for a variety of diseases. Previous work has suggested that peptide ligands bind to class B GPCRs according to a two-domain binding model, in which the C-terminal region of the peptide targets the ECD and the N-terminal region of the peptide binds to the TMD binding pocket. Recently, three structures of class B GPCRs in complex with peptide ligands have been solved. These structures provide essential insights into peptide ligand recognition by class B GPCRs. However, owing to resolution limitations, the specific molecular interactions for peptide binding to class B GPCRs remain ambiguous. Moreover, these previously solved structures have different ECD conformations relative to the TMD, which introduces questions regarding inter-domain conformational flexibility and the changes required for receptor activation. Here we report the 3.0 Å-resolution crystal structure of the full-length human glucagon receptor (GCGR) in complex with a glucagon analogue and partial agonist, NNC1702. This structure provides molecular details of the interactions between GCGR and the peptide ligand. It reveals a marked change in the relative orientation between the ECD and TMD of GCGR compared to the previously solved structure of the inactive GCGR-NNC0640-mAb1 complex. Notably, the stalk region and the first extracellular loop undergo major conformational changes in secondary structure during peptide binding, forming key interactions with the peptide. We further propose a dual-binding-site trigger model for GCGR activation-which requires conformational changes of the stalk, first extracellular loop and TMD-that extends our understanding of the previously established two-domain peptide-binding model of class B GPCRs.
Sujet(s)
Glucagon/analogues et dérivés , Récepteurs au glucagon/composition chimique , Récepteurs au glucagon/métabolisme , Cristallographie aux rayons X , Agonisme partiel des médicaments , Humains , Ligands , Modèles moléculaires , Liaison aux protéines , Conformation des protéinesRÉSUMÉ
The cannabinoid receptor 1 (CB1) is the principal target of the psychoactive constituent of marijuana, the partial agonist Δ9-tetrahydrocannabinol (Δ9-THC). Here we report two agonist-bound crystal structures of human CB1 in complex with a tetrahydrocannabinol (AM11542) and a hexahydrocannabinol (AM841) at 2.80 Å and 2.95 Å resolution, respectively. The two CB1-agonist complexes reveal important conformational changes in the overall structure, relative to the antagonist-bound state, including a 53% reduction in the volume of the ligand-binding pocket and an increase in the surface area of the G-protein-binding region. In addition, a 'twin toggle switch' of Phe2003.36 and Trp3566.48 (superscripts denote Ballesteros-Weinstein numbering) is experimentally observed and appears to be essential for receptor activation. The structures reveal important insights into the activation mechanism of CB1 and provide a molecular basis for predicting the binding modes of Δ9-THC, and endogenous and synthetic cannabinoids. The plasticity of the binding pocket of CB1 seems to be a common feature among certain class A G-protein-coupled receptors. These findings should inspire the design of chemically diverse ligands with distinct pharmacological properties.
Sujet(s)
Agonistes des récepteurs de cannabinoïdes/composition chimique , Dronabinol/analogues et dérivés , Dropéridol/analogues et dérivés , Récepteur cannabinoïde de type CB1/agonistes , Récepteur cannabinoïde de type CB1/composition chimique , Sites de fixation , Agonistes des récepteurs de cannabinoïdes/synthèse chimique , Agonistes des récepteurs de cannabinoïdes/pharmacologie , Cristallographie aux rayons X , Dronabinol/synthèse chimique , Dronabinol/composition chimique , Dronabinol/pharmacologie , Dropéridol/synthèse chimique , Dropéridol/composition chimique , Dropéridol/pharmacologie , Protéines G hétérotrimériques/métabolisme , Humains , Ligands , Simulation de docking moléculaire , Liaison aux protéines , Conformation des protéines , Récepteur cannabinoïde de type CB1/antagonistes et inhibiteurs , Récepteur cannabinoïde de type CB1/métabolismeRÉSUMÉ
Apelin receptor (APJR) is a key regulator of human cardiovascular function and is activated by two different endogenous peptide ligands, apelin and Elabela, each with different isoforms diversified by length and amino acid sequence. Here we report the 2.6-Å resolution crystal structure of human APJR in complex with a designed 17-amino-acid apelin mimetic peptide agonist. The structure reveals that the peptide agonist adopts a lactam constrained curved two-site ligand binding mode. Combined with mutation analysis and molecular dynamics simulations with apelin-13 binding to the wild-type APJR, this structure provides a mechanistic understanding of apelin recognition and binding specificity. Comparison of this structure with that of other peptide receptors suggests that endogenous peptide ligands with a high degree of conformational flexibility may bind and modulate the receptors via a similar two-site binding mechanism.
Sujet(s)
Récepteur de l'apeline/composition chimique , Alanine , Apeline/composition chimique , Récepteur de l'apeline/agonistes , Récepteur de l'apeline/génétique , Cristallographie aux rayons X , Humains , Simulation de dynamique moléculaire , Mimétisme moléculaire , Mutagenèse dirigée , Fragments peptidiques/composition chimique , Peptides cycliques/composition chimique , Conformation des protéines , Transduction du signalRÉSUMÉ
The glucagon-like peptide-1 receptor (GLP-1R) and the glucagon receptor (GCGR) are members of the secretin-like class B family of G-protein-coupled receptors (GPCRs) and have opposing physiological roles in insulin release and glucose homeostasis. The treatment of type 2 diabetes requires positive modulation of GLP-1R to inhibit glucagon secretion and stimulate insulin secretion in a glucose-dependent manner. Here we report crystal structures of the human GLP-1R transmembrane domain in complex with two different negative allosteric modulators, PF-06372222 and NNC0640, at 2.7 and 3.0 Å resolution, respectively. The structures reveal a common binding pocket for negative allosteric modulators, present in both GLP-1R and GCGR and located outside helices V-VII near the intracellular half of the receptor. The receptor is in an inactive conformation with compounds that restrict movement of the intracellular tip of helix VI, a movement that is generally associated with activation mechanisms in class A GPCRs. Molecular modelling and mutagenesis studies indicate that agonist positive allosteric modulators target the same general region, but in a distinct sub-pocket at the interface between helices V and VI, which may facilitate the formation of an intracellular binding site that enhances G-protein coupling.
Sujet(s)
Récepteur du peptide-1 similaire au glucagon/composition chimique , Récepteur du peptide-1 similaire au glucagon/métabolisme , Régulation allostérique/effets des médicaments et des substances chimiques , Site allostérique/effets des médicaments et des substances chimiques , Séquence d'acides aminés , Aminopyridines/composition chimique , Aminopyridines/métabolisme , Aminopyridines/pharmacologie , Benzamides/composition chimique , Benzamides/métabolisme , Benzamides/pharmacologie , Cristallographie aux rayons X , Récepteur du peptide-1 similaire au glucagon/agonistes , Humains , Modèles moléculaires , Phénylurées/composition chimique , Phénylurées/métabolisme , Phénylurées/pharmacologie , Domaines protéiquesRÉSUMÉ
The human glucagon receptor, GCGR, belongs to the class B G-protein-coupled receptor family and plays a key role in glucose homeostasis and the pathophysiology of type 2 diabetes. Here we report the 3.0 Å crystal structure of full-length GCGR containing both the extracellular domain and transmembrane domain in an inactive conformation. The two domains are connected by a 12-residue segment termed the stalk, which adopts a ß-strand conformation, instead of forming an α-helix as observed in the previously solved structure of the GCGR transmembrane domain. The first extracellular loop exhibits a ß-hairpin conformation and interacts with the stalk to form a compact ß-sheet structure. Hydrogen-deuterium exchange, disulfide crosslinking and molecular dynamics studies suggest that the stalk and the first extracellular loop have critical roles in modulating peptide ligand binding and receptor activation. These insights into the full-length GCGR structure deepen our understanding of the signalling mechanisms of class B G-protein-coupled receptors.
Sujet(s)
Récepteurs au glucagon/composition chimique , Récepteurs au glucagon/classification , Site allostérique/effets des médicaments et des substances chimiques , Benzamides/composition chimique , Benzamides/métabolisme , Benzamides/pharmacologie , Membrane cellulaire/métabolisme , Réactifs réticulants/composition chimique , Cristallographie aux rayons X , Mesure d'échange de deutérium , Disulfures/composition chimique , Humains , Ligands , Modèles moléculaires , Simulation de dynamique moléculaire , Phénylurées/composition chimique , Phénylurées/métabolisme , Phénylurées/pharmacologie , Domaines protéiques , Stabilité protéique , Récepteurs au glucagon/agonistes , Récepteurs au glucagon/métabolismeRÉSUMÉ
The Smoothened receptor (SMO) belongs to the Class Frizzled of the G protein-coupled receptor (GPCR) superfamily, constituting a key component of the Hedgehog signalling pathway. Here we report the crystal structure of the multi-domain human SMO, bound and stabilized by a designed tool ligand TC114, using an X-ray free-electron laser source at 2.9 Å. The structure reveals a precise arrangement of three distinct domains: a seven-transmembrane helices domain (TMD), a hinge domain (HD) and an intact extracellular cysteine-rich domain (CRD). This architecture enables allosteric interactions between the domains that are important for ligand recognition and receptor activation. By combining the structural data, molecular dynamics simulation, and hydrogen-deuterium-exchange analysis, we demonstrate that transmembrane helix VI, extracellular loop 3 and the HD play a central role in transmitting the signal employing a unique GPCR activation mechanism, distinct from other multi-domain GPCRs.
Sujet(s)
Protéines Hedgehog/métabolisme , Domaines protéiques , Transduction du signal , Récepteur Smoothened/composition chimique , Sites de fixation , Cristallographie aux rayons X , Mesure d'échange de deutérium/méthodes , Cellules HEK293 , Humains , Ligands , Spectrométrie de masse/méthodes , Simulation de dynamique moléculaire , Liaison aux protéines , Protéines recombinantes/composition chimique , Protéines recombinantes/isolement et purification , Protéines recombinantes/métabolisme , Récepteur Smoothened/isolement et purification , Récepteur Smoothened/métabolismeRÉSUMÉ
Cannabinoid receptor 1 (CB1) is the principal target of Δ9-tetrahydrocannabinol (THC), a psychoactive chemical from Cannabis sativa with a wide range of therapeutic applications and a long history of recreational use. CB1 is activated by endocannabinoids and is a promising therapeutic target for pain management, inflammation, obesity, and substance abuse disorders. Here, we present the 2.8 Å crystal structure of human CB1 in complex with AM6538, a stabilizing antagonist, synthesized and characterized for this structural study. The structure of the CB1-AM6538 complex reveals key features of the receptor and critical interactions for antagonist binding. In combination with functional studies and molecular modeling, the structure provides insight into the binding mode of naturally occurring CB1 ligands, such as THC, and synthetic cannabinoids. This enhances our understanding of the molecular basis for the physiological functions of CB1 and provides new opportunities for the design of next-generation CB1-targeting pharmaceuticals.
Sujet(s)
Antagonistes des récepteurs de cannabinoïdes/composition chimique , Morpholines/composition chimique , Pyrazoles/composition chimique , Récepteur cannabinoïde de type CB1/antagonistes et inhibiteurs , Récepteur cannabinoïde de type CB1/composition chimique , Sites de fixation , Cannabinoïdes/pharmacologie , Cannabis/composition chimique , Cristallographie aux rayons X , Dronabinol/pharmacologie , Endocannabinoïdes/pharmacologie , Humains , Ligands , Morpholines/synthèse chimique , Liaison aux protéines , Structure en hélice alpha , Pyrazoles/synthèse chimiqueRÉSUMÉ
The glucagon-like peptide-1 receptor (GLP-1R) belongs to the secretin-like (class B) family of G protein-coupled receptors. Members of the class B family are distinguished by their large extracellular domain, which works cooperatively with the canonical seven-transmembrane (7TM) helical domain to signal in response to binding of various peptide hormones. We have combined structure-based site-specific mutational studies with molecular dynamics simulations of a full-length model of GLP-1R bound to multiple peptide ligand variants. Despite the high sequence similarity between GLP-1R and its closest structural homologue, the glucagon receptor (GCGR), nearly half of the 62 stably expressed mutants affected GLP-1R in a different manner than the corresponding mutants in GCGR. The molecular dynamics simulations of wild-type and mutant GLP-1R·ligand complexes provided molecular insights into GLP-1R-specific recognition mechanisms for the N terminus of GLP-1 by residues in the 7TM pocket and explained how glucagon-mimicking GLP-1 mutants restored binding affinity for (GCGR-mimicking) GLP-1R mutants. Structural analysis of the simulations suggested that peptide ligand binding mode variations in the 7TM binding pocket are facilitated by movement of the extracellular domain relative to the 7TM bundle. These differences in binding modes may account for the pharmacological differences between GLP-1 peptide variants.
Sujet(s)
Glucagon-like peptide 1/métabolisme , Récepteur du peptide-1 similaire au glucagon/composition chimique , Récepteur du peptide-1 similaire au glucagon/métabolisme , Séquence d'acides aminés , Animaux , Sites de fixation , Cellules CHO , Cricetulus , Glucagon/métabolisme , Glucagon-like peptide 1/composition chimique , Glucagon-like peptide 1/génétique , Récepteur du peptide-1 similaire au glucagon/génétique , Cellules HEK293 , Humains , Simulation de dynamique moléculaire , Mutagenèse dirigée , Mutation ponctuelle , Liaison aux protéines , Structure tertiaire des protéinesRÉSUMÉ
Lipid biology continues to emerge as an area of significant therapeutic interest, particularly as the result of an enhanced understanding of the wealth of signaling molecules with diverse physiological properties. This growth in knowledge is epitomized by lysophosphatidic acid (LPA), which functions through interactions with at least six cognate G protein-coupled receptors. Herein, we present three crystal structures of LPA1 in complex with antagonist tool compounds selected and designed through structural and stability analyses. Structural analysis combined with molecular dynamics identified a basis for ligand access to the LPA1 binding pocket from the extracellular space contrasting with the proposed access for the sphingosine 1-phosphate receptor. Characteristics of the LPA1 binding pocket raise the possibility of promiscuous ligand recognition of phosphorylated endocannabinoids. Cell-based assays confirmed this hypothesis, linking the distinct receptor systems through metabolically related ligands with potential functional and therapeutic implications for treatment of disease.
Sujet(s)
Cristallographie aux rayons X , Sites de fixation , Chromatographie sur gel , Humains , Ligands , Modèles moléculaires , Récepteurs à l'acide phosphatidique/antagonistes et inhibiteurs , Récepteurs aux lysosphingolipides/composition chimique , Bibliothèques de petites moléculesRÉSUMÉ
The sphingosine 1 phosphate receptor family has been studied widely since the initial discovery of its first member, endothelium differentiation gene 1. Since this initial discovery, the family has been renamed and the primary member of the family, the S1P1 receptor, has been targeted for a variety of disease indications and successfully drugged for the treatment of patients with relapsing multiple sclerosis. Recently, the three-dimensional structure of the S1P1 receptor has been determined by X-ray crystallography and the specifics of the sphingosine 1 phosphate ligand binding pocket mapped. Key structural features for the S1P1 receptor will be reviewed and the potential binding modes of additional pharmacologically active agents against the receptor will be analyzed in an effort to better understand the structural basis of important receptor-ligand interactions.
Sujet(s)
Récepteurs aux lysosphingolipides/composition chimique , Séquence d'acides aminés , Animaux , Sites de fixation , Humains , Lysophospholipides/composition chimique , Lysophospholipides/métabolisme , Données de séquences moléculaires , Structure tertiaire des protéines , Récepteurs aux lysosphingolipides/génétique , Récepteurs aux lysosphingolipides/métabolisme , Sphingosine/analogues et dérivés , Sphingosine/composition chimique , Sphingosine/métabolismeRÉSUMÉ
Hewletts Creek, in Wilmington, North Carolina, drains a large suburban watershed and as such is affected by high fecal bacteria loads and periodic algal blooms from nutrient loading. During 2007, a 3.1-ha wetland was constructed to treat stormwater runoff from a 238-ha watershed within the Hewletts Creek drainage. A rain event sampling program was performed in 2009-2010 to evaluate the efficacy of the wetland in reducing pollutant loads from the stormwater runoff passing through the wetland. During the eight storms sampled, the wetland greatly moderated the hydrograph and retained and/or removed 50 to 75% of the inflowing stormwater volume. High removal rates of fecal coliform bacteria were achieved, with an average load reduction of 99% and overall concentration reduction of >90%. Particularly high (>90%) reductions of ammonium and orthophosphate loads also occurred, and lesser but still substantial reductions of total phosphorus (89%) and total suspended solids loads (88%) were achieved. Removal of nitrate was seasonally dependent, with lower removal occurring in cold weather and a high percentage (90%+) of nitrate load removal occurring in the growing season when water temperature exceeded 15°C. Long-term before-and-after sampling in downstream Hewletts Creek proper showed that, after wetland construction, statistically significant average decreases of 43% for nitrate, 72% for ammonium, and 59% for fecal coliform bacteria were realized. Wetland features contributing to the high pollutant control efficacy included available space for a large wetland, construction of deep forebays, and a dense and diverse aquatic and shoreline plant assemblage.