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1.
Structure ; 28(3): 371-377.e3, 2020 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-31978323

RESUMO

G-protein-coupled receptors (GPCRs) mediate a wide range of human physiological functions by transducing extracellular ligand binding events into intracellular responses. GPCRs can activate parallel, independent signaling pathways mediated by G proteins or ß-arrestins. Whereas "balanced" agonists activate both pathways equally, "biased" agonists dominantly activate one pathway, which is of interest for designing GPCR-targeting drugs because it may mitigate undesirable side effects. Previous studies demonstrated that ß-arrestin activation is associated with transmembrane helix VII (TM VII) of GPCRs. Here, single-molecule fluorescence spectroscopy with the ß2-adrenergic receptor (ß2AR) in the ligand-free state showed that TM VII spontaneously fluctuates between one inactive and one active-like conformation. The presence of the ß-arrestin-biased agonist isoetharine prolongs the dwell time of TM VII in the active-like conformation compared with the balanced agonist formoterol, suggesting that ligands can induce signaling bias by modulating the kinetics of receptor conformational exchange.

2.
Vet Pathol ; 57(1): 183-191, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31640487

RESUMO

Cardiovascular disorders and predominantly idiopathic myocardial fibrosis are frequently associated with mortality among zoo-housed chimpanzees (Pan troglodytes). Formalin-fixed whole hearts of deceased chimpanzees housed in zoos (n = 33) and an African sanctuary (n = 2) underwent detailed macroscopic and histopathologic examination using a standardized protocol. Archived histological slides from the hearts of 23 additional African sanctuary-housed chimpanzees were also examined. Myocardial fibrosis (MF) was identified in 30 of 33 (91%) of the zoo-housed chimpanzees but none of the 25 sanctuary-housed chimpanzees. MF was shown to be characterized by both interstitial and replacement fibrosis. Immunophenotyping demonstrated that the fibrotic lesions were accompanied by the increased presence of macrophages, alpha smooth muscle actin-positive myofibroblasts, and a minimal to mild T-cell-dominant leukocyte infiltration. There was no convincing evidence of cardiotropic viral infection or suggestion that diabetes mellitus or vitamin E or selenium deficiency were associated with the presence of the lesion. However, serum vitamin D concentrations among zoo-housed chimpanzees were found to be lower in seasons of low ultraviolet light levels.

3.
Int J Nurs Stud ; 102: 103467, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31810018

RESUMO

BACKGROUND: Cancer diagnosis and treatments may present severe adversity and challenge an individual adaptation ability. Individual resilience might be an important factor influencing the response to adversity in adult cancer patients. To date, the concept of individual resilience in adults with cancer has not been analysed in detail. OBJECTIVE: To conceptualise resilience in adult cancer care. DESIGN: A concept analysis. DATA SOURCES: Studies published between 2008 and 2019 were searched in MEDLINE, CINAHL, PsycINFO, Embase, Web of Knowledge and Scopus databases. Additional records were identified by manually searching reference lists of relevant studies. Quantitative and qualitative studies that investigated resilience in adult cancer care were included. METHODS: Walker and Avant's method of concept analysis was employed. The key elements for conceptualising resilience in adult cancer care (including defining attributes, antecedents and consequences of resilience) were identified from included articles. RESULTS: 53 included articles provided data for resilience conceptualisation. Analysis identified three defining attributes: (1) ego resiliency; (2) recalibration; and (3) interaction with the environment. Antecedents of resilience were cancer-related adversity, and consequences of resilience which were divided into two categories: (1) psychological distress level; and (2) individual functioning performance (physical, psychological and social functioning). CONCLUSIONS: Our conceptualisation of individual resilience based on the defining attributes, antecedents and consequences resulted in a preliminary conceptual model. This model can be further tested and may inform the development of cancer-specific resilience measures and interventions for adults with cancer focusing on individual resilience.

4.
J Feline Med Surg ; 22(2): 77-83, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-30719951

RESUMO

OBJECTIVES: The aim of this study was to compare the quality of anaesthesia and analgesia between methadone and buprenorphine in combination with medetomidine after induction with intramuscular (IM) alfaxalone in cats undergoing ovariohysterectomy. METHODS: Fifty-one female cats (American Society of Anesthesiologists status I-II), with a median age of 12 months (range 2-60 months), weighing a mean ± SD of 2.5 ± 0.5 kg, were recruited to the study. Cats were randomly allocated to receive medetomidine (600 µg/m2) and buprenorphine (180 µg/m2) (group MB) or medetomidine (500 µg/m2) and methadone (5 mg/m2) (group MM) IM. Anaesthesia was induced 15 mins later using alfaxalone (3 mg/kg) IM. Anaesthesia was maintained with isoflurane in oxygen. All cats received meloxicam preoperatively. Quality of premedication and induction and intraoperative physiological parameters were recorded. Atipamezole (50% of medetomidine dose) was administered at the end of surgery. Cats were assessed postoperatively by the same blinded observer using a simple descriptive scale, numeric rating scale, dynamic interactive visual analogue scale (DIVAS) and UNESP-Botucatu multidimensional composite pain scales, at 10, 20 and 30 mins post-extubation. Parametric and non-parametric data were compared using Student's t-test or Mann-Whitney U-tests, respectively. RESULTS: Forty-one cats completed the study. No significant differences were detected between groups before or during anaesthesia. No cats required rescue analgesia. DIVAS scores at 10 mins were significantly less in the MM group compared with the MB. No differences between groups at any other time points were detected using the four metrology instruments. CONCLUSIONS AND RELEVANCE: Both protocols provided good anaesthesia conditions for ovariohysterectomy in the cat.

5.
Sci Adv ; 5(11): eaax9115, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31807708

RESUMO

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.

6.
Structure ; 27(12): 1745-1759, 2019 12 03.
Artigo em Inglês | MEDLINE | ID: mdl-31780431

RESUMO

Structures of biomolecular systems are increasingly computed by integrative modeling. In this approach, a structural model is constructed by combining information from multiple sources, including varied experimental methods and prior models. In 2019, a Workshop was held as a Biophysical Society Satellite Meeting to assess progress and discuss further requirements for archiving integrative structures. The primary goal of the Workshop was to build consensus for addressing the challenges involved in creating common data standards, building methods for federated data exchange, and developing mechanisms for validating integrative structures. The summary of the Workshop and the recommendations that emerged are presented here.

7.
HPB (Oxford) ; 2019 Oct 18.
Artigo em Inglês | MEDLINE | ID: mdl-31635959

RESUMO

BACKGROUND: The aim of this systematic review is to examine patient-reported outcome measures (PROMs), their attributes and application in patients with pancreatic cancer (PC). METHOD: A systematic literature search was undertaken of articles published to June 2018 to identify PROMs applied in primary studies in PC. Characteristics of the included studies and PROMs were described with identified scales grouped into five domains. The psychometric properties of the identified PROMs were further assessed for reliability and validity among patients with PC. RESULTS: From 1688 studies screened, 170 were included. Almost half (48%) were conducted in patients with unresectable PC; the majority of these (68%) were evaluated in randomized controlled trials. Median questionnaire completion rates fell below 10% of the original cohort within 12 months in patients with unresectable PC compared to 75% in patients with resectable PC. Seventy PROMs were identified, 32 measuring unidimensional parameters (e.g. pain) and 35 measuring multidimensional (e.g. quality of life) constructs. Only five (7%) PROMs were disease-specific and 13 (19%) were validated in patients with PC. Fifty scales were grouped into 19 physical, 9 psychological, 6 psychiatric, 9 social and 7 other domains. CONCLUSION: Three multidimensional PROMs, the: (i) FACT-HEP in unresectable PC; (ii) QLQ-PAN26 (in conjunction with its core QLQ-C30 PROM) in resectable PC; and (iii) MDASI-GI are recommended as instruments to capture quality of life in patients with PC. Summarised scales and psychometric evaluation provide a framework to choose PROMs for scales not captured by the recommended PROMs.

8.
J Neurosci ; 39(48): 9546-9559, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31628176

RESUMO

Sensory cortices process stimuli in manners essential for perception. Very little is known regarding interactions between olfactory cortices. The piriform "primary" olfactory cortex, especially its anterior division (aPCX), extends dense association fibers into the ventral striatum's olfactory tubercle (OT), yet whether this corticostriatal pathway is capable of shaping OT activity, including odor-evoked activity, is unknown. Further unresolved is the synaptic circuitry and the spatial localization of OT-innervating PCX neurons. Here we build upon standing literature to provide some answers to these questions through studies in mice of both sexes. First, we recorded the activity of OT neurons in awake mice while optically stimulating principal neurons in the aPCX and/or their association fibers in the OT while the mice were delivered odors. This uncovered evidence that PCX input indeed influences OT unit activity. We then used patch-clamp recordings and viral tracing to determine the connectivity of aPCX neurons upon OT neurons expressing dopamine receptor types D1 or D2, two prominent cell populations in the OT. These investigations uncovered that both populations of neurons receive monosynaptic inputs from aPCX glutamatergic neurons. Interestingly, this input originates largely from the ventrocaudal aPCX. These results shed light on some of the basic physiological properties of this pathway and the cell-types involved and provide a foundation for future studies to identify, among other things, whether this pathway has implications for perception.SIGNIFICANCE STATEMENT Sensory cortices interact to process stimuli in manners considered essential for perception. Very little is known regarding interactions between olfactory cortices. The present study sheds light on some of the basic physiological properties of a particular intercortical pathway in the olfactory system and provides a foundation for future studies to identify, among other things, whether this pathway has implications for perception.

9.
Qual Life Res ; 28(11): 2871-2887, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31364037

RESUMO

INTRODUCTION: The prognostic value of patient-reported outcomes (PROs) has been determined in some cancers, but a focussed review in colorectal cancer (CRC) has not yet been conducted. We systematically reviewed PRO predictors of CRC patient survival. METHODS: We searched four electronic databases (from inception to May 2018), reference lists and professional organisations to identify studies reporting pre-treatment PRO predictors of overall survival (OS) or progression-free survival (PFS) in CRC identified through univariate or multivariate models. Two reviewers independently applied inclusion criteria and extracted data on study characteristics, median and 1-year survival rates, PROs assessed and model results. RESULTS: In 25 of 27 studies (n = 12,544), at least one PRO was significantly associated with survival. Physical functioning, fatigue, pain and appetite loss predicted OS more often than other PROs in metastatic disease (19/27 studies). One study explored PRO predictors in early-stage CRC, finding emotional well-being and mood predicted OS. In mixed-stage samples (7/27 studies), physical functioning predicted OS more often than other PROs. Few studies modelled PFS, for which few PROs had predictive value. CONCLUSIONS: Physical and psychological functioning, pain, fatigue and appetite loss had prognostic significance above and beyond clinical predictors in CRC. Routine monitoring of these PROs may allow earlier detection and amelioration of problems, which may improve quality of life and perhaps extend survival. More research is needed to determine prognostic value of PROs in early-stage CRC, and prognostic significance of changes in PRO scores.


Assuntos
Neoplasias Colorretais/mortalidade , Medidas de Resultados Relatados pelo Paciente , Qualidade de Vida/psicologia , Neoplasias Colorretais/tratamento farmacológico , Feminino , Humanos , Masculino , Prognóstico
10.
Chem Commun (Camb) ; 55(40): 5724-5727, 2019 May 14.
Artigo em Inglês | MEDLINE | ID: mdl-31038495

RESUMO

Giant unilamellar protein vesicles (GUPs) were formed with the adenosine A2A receptor (A2AR) incorporated in the lipid bilayer and protein partitioning into the liquid ordered and liquid disordered phases was observed. When no ligand is bound, A2AR partitions preferentially into the liquid disordered phase of GUPs, while ligand-bound A2AR partitions into the liquid ordered phase.


Assuntos
Lipídeos/química , Receptores A2 de Adenosina/metabolismo , Animais , Sítios de Ligação , Células HEK293 , Humanos , Ligantes , Bicamadas Lipídicas , Transição de Fase , Ligação Proteica , Células Sf9
11.
Australas J Ageing ; 2019 May 09.
Artigo em Inglês | MEDLINE | ID: mdl-31069919

RESUMO

OBJECTIVE: To gain insights into the experience and challenges faced by Australasian geriatricians who have recently made the transition from advanced trainee to consultant. METHODS: An interpretative exploratory qualitative study. Geriatricians with five or less years of experience as consultants were recruited by email. Data were collected through semi-structured interviews, with themes identified through open axial coding. RESULTS: Respondents (n = 20) experienced a transition period in which they adjusted to the roles of final decision-maker and manager. Respondents felt relatively confident with their clinical skills, but under-prepared for non-clinical roles associated with becoming a consultant. Most respondents described challenges with career planning. Support networks were considered critical. CONCLUSIONS: This is the first study in Australasia exploring the transition from trainee to consultant geriatrician. Training programs should endeavour to create "consultant-like roles" during advanced training and address non-clinical competencies. Participants perceived that there should be more emphasis on career planning and mentorship.

12.
Psychooncology ; 28(5): 1048-1055, 2019 05.
Artigo em Inglês | MEDLINE | ID: mdl-30828923

RESUMO

OBJECTIVES: This study aimed to describe the levels of health literacy and experience of care coordination among Chinese migrant patients with cancer and their carers in Australia, and to examine factors associated with these. METHODS: Patients' self-reported data were collected using the Health Literacy and Cancer Care Coordination questionnaires. We conducted multivariate linear regression analyses to investigate predictors of patients' health literacy and their care experience. Canonical correlation analysis was used to examine the relationship between patients' health literacy and their care experience. RESULTS: A total of 68 patients and eight carers participated in the survey. Patients and carers reported similar levels of health literacy, with the lowest scores being in the "Having sufficient information to manage health" and "Navigating the health system" subscales. Gender (P = 0.026, partial η2  = 0.281) and educational attainment (P = 0.015, partial η2  = 0.250) had significant and large effects on patients' health literacy, after controlling for each other. Educational attainment showed a significant and medium association with patients' experience of cancer care coordination (P = 0.041, partial η2  = 0.101). A large and positive correlation was found between patients' health literacy and experience of cancer care coordination (canonical correlation = 0.81). CONCLUSIONS: Our findings reveal the health literacy and care coordination needs of Chinese migrant patients with cancer in Australia, especially those with lower educational attainment. Future efforts are necessary to enhance Chinese migrants' health literacy and establish an accessible and easy-to-navigate care environment.

13.
BMC Med ; 17(1): 58, 2019 03 12.
Artigo em Inglês | MEDLINE | ID: mdl-30857521

RESUMO

BACKGROUND: Between August and December 2017, more than 625,000 Rohingya from Myanmar fled into Bangladesh, settling in informal makeshift camps in Cox's Bazar district and joining 212,000 Rohingya already present. In early November, a diphtheria outbreak hit the camps, with 440 reported cases during the first month. A rise in cases during early December led to a collaboration between teams from Médecins sans Frontières-who were running a provisional diphtheria treatment centre-and the London School of Hygiene and Tropical Medicine with the goal to use transmission dynamic models to forecast the potential scale of the outbreak and the resulting resource needs. METHODS: We first adjusted for delays between symptom onset and case presentation using the observed distribution of reporting delays from previously reported cases. We then fit a compartmental transmission model to the adjusted incidence stratified by age group and location. Model forecasts with a lead time of 2 weeks were issued on 12, 20, 26 and 30 December and communicated to decision-makers. RESULTS: The first forecast estimated that the outbreak would peak on 19 December in Balukhali camp with 303 (95% posterior predictive interval 122-599) cases and would continue to grow in Kutupalong camp, requiring a bed capacity of 316 (95% posterior predictive interval (PPI) 197-499). On 19 December, a total of 54 cases were reported, lower than forecasted. Subsequent forecasts were more accurate: on 20 December, we predicted a total of 912 cases (95% PPI 367-2183) and 136 (95% PPI 55-327) hospitalizations until the end of the year, with 616 cases actually reported during this period. CONCLUSIONS: Real-time modelling enabled feedback of key information about the potential scale of the epidemic, resource needs and mechanisms of transmission to decision-makers at a time when this information was largely unknown. By 20 December, the model generated reliable forecasts and helped support decision-making on operational aspects of the outbreak response, such as hospital bed and staff needs, and with advocacy for control measures. Although modelling is only one component of the evidence base for decision-making in outbreak situations, suitable analysis and forecasting techniques can be used to gain insights into an ongoing outbreak.


Assuntos
Difteria/fisiopatologia , Surtos de Doenças/estatística & dados numéricos , Bangladesh , Humanos , Mianmar
14.
HPB (Oxford) ; 21(4): 444-455, 2019 04.
Artigo em Inglês | MEDLINE | ID: mdl-30316625

RESUMO

BACKGROUND: Best practise care optimises survival and quality of life in patients with pancreatic cancer (PC), but there is evidence of variability in management and suboptimal care for some patients. Monitoring practise is necessary to underpin improvement initiatives. We aimed to develop a core set of quality indicators that measure quality of care across the disease trajectory. METHODS: A modified, three-round Delphi survey was performed among experts with wide experience in PC care across three states in Australia. A total of 107 potential quality indicators were identified from the literature and divided into five areas: diagnosis and staging, surgery, other treatment, patient management and outcomes. A further six indicators were added by the panel, increasing potential quality indicators to 113. Rated on a scale of 1-9, indicators with high median importance and feasibility (score 7-9) and low disagreement (<1) were considered in the candidate set. RESULTS: From 113 potential quality indicators, 34 indicators met the inclusion criteria and 27 (7 diagnosis and staging, 5 surgical, 4 other treatment, 5 patient management, 6 outcome) were included in the final set. CONCLUSIONS: The developed indicator set can be applied as a tool for internal quality improvement, comparative quality reporting, public reporting and research in PC care.

15.
Nat Chem Biol ; 15(2): 206, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30573766

RESUMO

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.

16.
Nat Chem Biol ; 15(1): 11-17, 2019 01.
Artigo em Inglês | MEDLINE | ID: mdl-30510194

RESUMO

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.


Assuntos
Misoprostol/química , Receptores de Prostaglandina E Subtipo EP3/química , Receptores de Prostaglandina E Subtipo EP3/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Dinoprostona/análogos & derivados , Dinoprostona/química , Dinoprostona/metabolismo , Humanos , Misoprostol/metabolismo , Simulação de Acoplamento Molecular , Mutagênese Sítio-Dirigida , Conformação Proteica , Receptores de Prostaglandina E Subtipo EP3/agonistas , Receptores de Prostaglandina E Subtipo EP3/genética , Transdução de Sinais , Água/química
17.
Vet Anaesth Analg ; 45(6): 794-801, 2018 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-30314798

RESUMO

OBJECTIVE: To characterize the cardiopulmonary characteristics of two different anaesthetic protocols (tiletamine/zolazepam ± medetomidine) and their suitability for the immobilization of healthy chimpanzees undergoing cardiac assessment. STUDY DESIGN: Prospective, clinical, longitudinal study. ANIMALS: Six chimpanzees (Pan troglodytes) aged 4-16 years weighing 19.5-78.5 kg were anaesthetized on two occasions. METHODS: Anaesthesia was induced with tiletamine/zolazepam (TZ) (3-4 mg kg-1) or tiletamine/zolazepam (2 mg kg-1) and medetomidine (0.02 mg kg-1) (TZM) via blow dart [intramuscular (IM)] and maintained with intermittent boluses of ketamine (IV) or zolazepam/tiletamine (IM) as required. The overall quality of the anaesthesia was quantified based on scores given for: quality of induction, degree of muscle relaxation and ease of intubation. The time to achieve a light plane of anaesthesia, number of supplemental boluses needed and recovery characteristics were also recorded. Chimpanzees were continuously monitored and heart rate (HR), pulse rate (PR), respiratory rate (fR) oxygen saturation of haemoglobin (SpO2), systolic arterial pressure (SAP), diastolic arterial pressure (DAP), mean arterial pressure (MAP), rectal temperature, mucous membrane colour and capillary refill time recorded. During the first procedure (TZ) animals underwent a 12-channel electrocardiogram (ECG), haematology, biochemistry and cardiac biomarker assessment to rule out the presence of pre-existing cardiovascular disease. A detailed echocardiographic examination was carried out by the same blinded observer during both procedures. Data were compared using Student's paired t-test or Wilcoxon rank tests as appropriate. RESULTS: There was a significant difference for the area under the curves between anaesthetic protocols for HR, SAP, MAP and fR. No significant differences in the echocardiographic measurements were evident. Quality of anaesthesia was significantly better with TZM and no additional boluses were required. The TZ protocol required multiple supplemental boluses. CONCLUSIONS AND CLINICAL RELEVANCE: Both combinations are suitable for immobilization and cardiovascular evaluation of healthy chimpanzees. Further work is required to evaluate the effect of medetomidine in cardiovascular disease.


Assuntos
Anestésicos/farmacologia , Hemodinâmica/efeitos dos fármacos , Medetomidina/farmacologia , Respiração/efeitos dos fármacos , Tiletamina/farmacologia , Zolazepam/farmacologia , Anestésicos Combinados/farmacologia , Animais , Protocolos Clínicos , Combinação de Medicamentos , Feminino , Estudos Longitudinais , Masculino , Pan troglodytes , Estudos Prospectivos
18.
Cell ; 173(1): 11-19, 2018 03 22.
Artigo em Inglês | MEDLINE | ID: mdl-29570991

RESUMO

The construction of a predictive model of an entire eukaryotic cell that describes its dynamic structure from atomic to cellular scales is a grand challenge at the intersection of biology, chemistry, physics, and computer science. Having such a model will open new dimensions in biological research and accelerate healthcare advancements. Developing the necessary experimental and modeling methods presents abundant opportunities for a community effort to realize this goal. Here, we present a vision for creation of a spatiotemporal multi-scale model of the pancreatic ß-cell, a relevant target for understanding and modulating the pathogenesis of diabetes.


Assuntos
Células Secretoras de Insulina/metabolismo , Modelos Biológicos , Biologia Computacional , Descoberta de Drogas , Humanos , Células Secretoras de Insulina/citologia , Proteínas/química , Proteínas/metabolismo
19.
Structure ; 26(2): 259-269.e5, 2018 02 06.
Artigo em Inglês | MEDLINE | ID: mdl-29395784

RESUMO

Sodium ions are endogenous allosteric modulators of many G-protein-coupled receptors (GPCRs). Mutation of key residues in the sodium binding motif causes a striking effect on G-protein signaling. We report the crystal structures of agonist complexes for two variants in the first sodium coordination shell of the human A2A adenosine receptor, D522.50N and S913.39A. Both structures present an overall active-like conformation; however, the variants show key changes in the activation motif NPxxY. Changes in the hydrogen bonding network in this microswitch suggest a possible mechanism for modified G-protein signaling and enhanced thermal stability. These structures, signaling data, and thermal stability analysis with a panel of pharmacological ligands provide a basis for understanding the role of the sodium-coordinating residues on stability and G-protein signaling. Utilizing the D2.50N variant is a promising method for stabilizing class A GPCRs to accelerate structural efforts and drug discovery.


Assuntos
Regulação Alostérica/fisiologia , Sítio Alostérico/fisiologia , Receptores Acoplados a Proteínas-G/metabolismo , Transdução de Sinais/fisiologia , Humanos , Ligação Proteica , Conformação Proteica
20.
Cell ; 172(1-2): 68-80.e12, 2018 01 11.
Artigo em Inglês | MEDLINE | ID: mdl-29290469

RESUMO

Signaling across cellular membranes, the 826 human G protein-coupled receptors (GPCRs) govern a wide range of vital physiological processes, making GPCRs prominent drug targets. X-ray crystallography provided GPCR molecular architectures, which also revealed the need for additional structural dynamics data to support drug development. Here, nuclear magnetic resonance (NMR) spectroscopy with the wild-type-like A2A adenosine receptor (A2AAR) in solution provides a comprehensive characterization of signaling-related structural dynamics. All six tryptophan indole and eight glycine backbone 15N-1H NMR signals in A2AAR were individually assigned. These NMR probes provided insight into the role of Asp522.50 as an allosteric link between the orthosteric drug binding site and the intracellular signaling surface, revealing strong interactions with the toggle switch Trp 2466.48, and delineated the structural response to variable efficacy of bound drugs across A2AAR. The present data support GPCR signaling based on dynamic interactions between two semi-independent subdomains connected by an allosteric switch at Asp522.50.


Assuntos
Regulação Alostérica , Receptor A2A de Adenosina/química , Transdução de Sinais , Agonistas do Receptor A2 de Adenosina/química , Agonistas do Receptor A2 de Adenosina/farmacologia , Sítio Alostérico , Animais , Simulação de Acoplamento Molecular , Pichia , Ligação Proteica , Receptor A2A de Adenosina/metabolismo , Células Sf9 , Spodoptera
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