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1.
Cell ; 184(4): 931-942.e18, 2021 Feb 18.
Artigo em Inglês | MEDLINE | ID: mdl-33571431

RESUMO

The D1- and D2-dopamine receptors (D1R and D2R), which signal through Gs and Gi, respectively, represent the principal stimulatory and inhibitory dopamine receptors in the central nervous system. D1R and D2R also represent the main therapeutic targets for Parkinson's disease, schizophrenia, and many other neuropsychiatric disorders, and insight into their signaling is essential for understanding both therapeutic and side effects of dopaminergic drugs. Here, we report four cryoelectron microscopy (cryo-EM) structures of D1R-Gs and D2R-Gi signaling complexes with selective and non-selective dopamine agonists, including two currently used anti-Parkinson's disease drugs, apomorphine and bromocriptine. These structures, together with mutagenesis studies, reveal the conserved binding mode of dopamine agonists, the unique pocket topology underlying ligand selectivity, the conformational changes in receptor activation, and potential structural determinants for G protein-coupling selectivity. These results provide both a molecular understanding of dopamine signaling and multiple structural templates for drug design targeting the dopaminergic system.

2.
Acta Cir Bras ; 36(1): e360102, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33605307

RESUMO

PURPOSE: To study the Periplaneta americana L. extract Ento-B on the treatment of chronic ulcerative colitis induced by 2,4-dinitrochlorobenzene and acetic acid in rats and to explore its primary mechanism of action. METHODS: Using 2,4-dinitrochlorobenzene combined with acetic acid to induce chronic ulcerative colitis (chronic UC) in rats. The sulfasalazine (400 mg/kg) and Ento-B (200 mg/kg, 100 mg/kg,50 mg/kg) were given by intragastric administration and the effect was evaluated according to the disease activity index (DAI) score, colon mucosal injury index (CMDI) score, histopathological score (HS) and the serum levels of Interleukin-4(IL-4), Interleukin-10(IL-10), Tumor necrosis factor-α(TNF-α), Malondialdehyde(MDA), Superoxide dismutase(SOD) and Inducible nitric oxide synthase(iNOS.). RESULTS: Compared with the model group, all doses of Ento-B could reduce the score of CMDI (p < 0.05), HS(p < 0.05 or p < 0.01), significantly increased the expression of IL-4, IL-10, SOD (p < 0.01) and decreased the levels of TNF-α, MDA, iNOS in serum of UC rats, significantly improving the degree of colon lesionsin UC rats. CONCLUSIONS: Ento-B may play an important role in the treatment of ulcerative colitis induced byUC rats. The mechanism may be related to the increased expression of IL-4, IL-10, SOD and reduced expression of TNF-α, MDA, iNOS.


Assuntos
Colite Ulcerativa , Periplaneta , Ácido Acético , Animais , Colite Ulcerativa/induzido quimicamente , Colite Ulcerativa/tratamento farmacológico , Colo , Dinitroclorobenzeno , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Ratos , Fator de Necrose Tumoral alfa
3.
Cell Chem Biol ; 2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33545070

RESUMO

As a typical member of intrinsically disordered proteins (IDPs), HMGA1a carries many post-translational modifications (PTMs). To study the undefined function of acidic tail phosphorylations, seven HMGA1a proteins with site-specific modification(s) were chemically synthesized via Ser/Thr ligation. We found that the phosphorylations significantly inhibit HMGA1a-P53 interaction and the phosphorylations can induce conformational change of HMGA1a from an "open state" to a "close state." Notably, the positively charged lysine-arginine (KR) clusters are responsible for modulating HMGA1a conformation via electrostatic interaction with the phosphorylated acidic tail. Finally, we used a synthetic protein-affinity purification mass spectrometry (SP-AP-MS) methodology to profile the specific interactors, which further supported the function of HMGA1a phosphorylation. Collectively, this study highlights a mechanism for regulating IDPs' conformation and function by phosphorylation of non-protein-binding domain and showcases that the protein chemical synthesis in combination with mass spectrometry can serve as an efficient tool to study the IDPs' PTMs.

4.
Nanoscale ; 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33565548

RESUMO

Thin-film transistors (TFTs) have been widely used in the increasingly advanced field of displays. However, it remains a challenge for TFTs to overcome the poor subthreshold swing in the fast switching and high-speed applications. Here, we provide a solution to the above-mentioned challenge via supercritical dehydroxylation, which combines a low temperature, environmentally friendly supercritical fluid technology with a CaCl2 treatment. An embedded structure of amorphous indium gallium zinc oxide (a-IGZO) TFTs with double-layer high-k dielectric containing Ta2O5 and SiO2 layers was first manufactured. The subthreshold swing of the fabricated TFTs treated with supercritical dehydroxylation was optimized to an ultra-low value of 72.7 mV dec-1. Moreover, other key figures of merits including threshold voltage, on/off ratio and field effect mobility all improved after the supercritical dehydroxylation. The bandgap of the gate dielectric material increased due to the supercritical dehydroxylation verified by the current conduction mechanism. Besides, numerous material analyses further confirmed that owing to the supercritical dehydroxylation the dominant dehydration reactions can effectively repair the defects introduced in the device manufacture. The ultra-low subthreshold swing with optimized electrical performances can be achieved via the low-temperature supercritical dehydroxylation treatment, enabling its promising potential in realizing ultra-fast and low power electronics.

5.
Mil Med ; 2021 Feb 12.
Artigo em Inglês | MEDLINE | ID: mdl-33576405

RESUMO

BACKGROUND: External hemorrhage control devices (EHCDs) are effective in reducing the death risk of noncompressible torso hemorrhage (NCTH), but the pressurized area is too large to prevent serious organ damage. This study aims to establish the surface localization strategy of EHCDs based on the anatomical features of NCTH-related arteries through CT images to facilitate the optimal design and application of EHCDs. METHODS: Two hundred patients who underwent abdominal CT were enrolled. Anatomical parameters such as the length of the common iliac artery (CIA), the external iliac artery (EIA), and the common femoral artery were measured; positional relationships among the EHCD-targeted arteries, umbilicus, anterior superior iliac spine (ASIS), and pubic tubercle (PT) were determined. The accuracy of surface localization was verified by the 3D-printed mannequins of 20 real patients. RESULTS: Aortic bifurcation (AB) was 7.5 ± 8.6 mm to the left of the umbilicus. The left CIA (left: 46.6 ± 16.0 mm vs. right: 43.3 ± 15.5 mm, P = .038) and the right EIA (left: 102.6 ± 16.3 mm vs. right: 111.5 ± 18.8 mm, P < .001) were longer than their counterparts, respectively. The vertical distance between the CIA terminus and the ipsilateral AB-ASIS line was 19.6 ± 8.2 mm, and the left and right perpendicular intersections were located at the upper one-third and one-fourth of the AB-ASIS line, respectively. The length ratio of EIA-ASIS to ASIS-PT was 0.6:1. The predicted point and its actual subpoint were significantly correlated (P ≤ .002), and the vertical distance between the two points was ≤5.5 mm. CONCLUSION: The arterial localization strategy established via anatomical investigation was consistent with the actual situation. The data are necessary for improving EHCD design, precise hemostasis, and EHCD-related collateral injuries.Trial registration: Ratification no. 2019092. Registered November 4, 2020-retrospectively registered, www.chictr.org.cn.

6.
J Orthop Surg Res ; 16(1): 30, 2021 Jan 09.
Artigo em Inglês | MEDLINE | ID: mdl-33422082

RESUMO

PURPOSE: D-dimer and fibrinogen, both belonging to coagulation parameters, are controversial for the diagnosis of periprosthetic joint infection (PJI). This meta-analysis was conducted to compare their diagnostic accuracies for PJI by synthesizing currently available evidence. METHODS: Cochrane Library, MEDLINE, Web of Science, and Embase up to March 1, 2020, and other relevant articles were searched. Five hundred and eighty-one articles were identified after initial research, and 11 studies were included finally. No threshold effects were found between studies. The pooled sensitivity, specificity, and positive and negative likelihood ratio were reported to evaluate the diagnostic performance with heterogeneity analysis. Z test statistics was used to analyze the difference of diagnostic performance between D-dimer and fibrinogen. RESULTS: The pooled sensitivity, specificity, and positive and negative likelihood ratio of D-dimer for PJI were 0.79 (95% [CI], 0.72-0.85), 0.77 (0.67-0.84), 3.38 (2.21-5.18), and 0.27 (0.18-0.41), respectively. As for fibrinogen, the pooled sensitivity, specificity, and positive and negative likelihood ratio for PJI were 0.75 (0.68-0.80), 0.85 (0.82-0.88), 5.12 (4.22-6.22), and 0.30 (0.23-0.37), respectively. Great heterogeneity was found in studies for D-dimer, and univariate meta-regression analysis revealed that number of involved joints, disease spectrum, comorbidities influencing D-dimer, and sample sources were the source of heterogeneity. Z test found that the pooled specificity of fibrinogen was significantly higher than D-dimer (0.85 ± 0.01 versus 0.77 ± 0.04, p = 0.03). The pooled positive likelihood ratio of fibrinogen was significantly higher than D-dimer (5.12 ± 0.51 versus 3.38 ± 0.74, p = 0.03). CONCLUSION: Based on currently available evidence, the meta-analysis suggests that fibrinogen performs better than D-dimer as a rule-in diagnostic tool for its higher specificity. However, more prospective trials with larger size are still needed to provide further confirmation. TRIAL REGISTRATION: This meta-analysis was prospectively registered on PROSPERO (International prospective register of systematic reviews), and the registering number was CRD42020177176 .

7.
Inorg Chem ; 2021 Jan 27.
Artigo em Inglês | MEDLINE | ID: mdl-33501824

RESUMO

Increasing the thermostability of α-diimine late-transition-metal complexes and therefore rendering them more active at higher temperatures is of great importance, yet challenging for the olefin polymerization field. In the present research, a new family of α-diimine palladium complexes that can promote norbornene polymerization at high temperatures (up to 140 °C) is disclosed. Because of the conformational restriction caused by increasing the axial and equatorial bulkiness as well as the presence of intraligand H···F hydrogen bonds, N-aryl rotations can be efficiently restricted, therefore circumventing the deactivation of the active species at high temperatures. At 80-140 °C, these complexes can efficiently catalyze norbornene homopolymerizations, giving high catalytic activities up to 5.65 × 107 g of PNB per mole Ni per hour and polymers with high molecular weights up to 37.2 × 104 g/mol, which are highly superior to catalytic systems mediated by CF3-free complexes. Moreover, these complexes could also afford medium catalytic activities in the presence of polar 5-norbornene-2-carboxylic acid methyl ester (NB-COOCH3).

8.
BMC Infect Dis ; 21(1): 83, 2021 Jan 19.
Artigo em Inglês | MEDLINE | ID: mdl-33468062

RESUMO

BACKGROUND: Hepatitis B virus (HBV) infection is a major concern for blood safety in high-prevalence HBV countries such as China. In Shenzhen, dual hepatitis B surface antigen (HBsAg) enzyme-linked immunosorbent assays (ELISAs) have been adopted in parallel with nucleic acid testing (NAT) for donors for over a decade. A small proportion of blood donors test reactive (R) for HBsAg but negative through routine NAT, which can lead to HBV infection with an extremely low viral load. OBJECTIVES: We aimed to investigate and analyze the molecular characteristics of HBV among blood donors that tested HBsAg R in a single ELISA test. METHODS: Blood donations were evaluated in this study if confirmed HBsAg R through one of two ELISA kits. Samples with non-reactive (NR) results by NAT were collected and tested for HBsAg by chemiluminescent microparticle immunoassay (CLIA) with a neutralization test. The level of HBsAg was further assessed by electrochemiluminescence immunoassay (ECLIA). The viral basic core promoter (BCP) and pre-core (PC) and S regions were amplified by nested PCR. Quantitative real-time PCR (qPCR) for viral load determination and individual donation (ID)-NAT were adopted simultaneously. HBsAg was confirmed with CLIA, ECLIA, nested PCR, qPCR, and ID-NAT. RESULTS: Of the 100,252 donations, 38 and 41 were identified as HBsAg R with Wantai and DiaSorin ELISA kits, respectively. Seventy-nine (0.077%, 79/100,252) blood samples with ELISA R-NR and NAT NR results were enrolled in the study. Of these, 17 (21.5%,17/79) were confirmed as HBsAg-positive. Of the 14 genotyped cases, 78.6% (11/14) were genotype B, and C and D were observed in two and one sample, respectively. Mutations were found in the S gene, including Y100C, Y103I, G145R, and L175S, which can affect the detection of HBsAg. A high-frequency mutation, T1719G (93.3%), was detected in the BCP/PC region, which reduced the viral replication. CONCLUSION: A small number of blood samples with HBsAg ELISA R-NR and NAT NR results were confirmed as HBV infection, viral nucleic acids were found in most of the samples through routine NAT methods. It is necessary to employ more sensitive and specific assays for the detection of HBV infection among blood donors.


Assuntos
Doadores de Sangue , Antígenos de Superfície da Hepatite B/sangue , Vírus da Hepatite B/genética , Hepatite B , China , DNA Viral/genética , Ensaio de Imunoadsorção Enzimática , Humanos , Técnicas de Amplificação de Ácido Nucleico , Filogenia , Reação em Cadeia da Polimerase/métodos
9.
Cytotherapy ; 23(3): 211-222, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33334686

RESUMO

BACKGROUND AIMS: Chimeric antigen receptor (CAR) T-cell therapy is a promising treatment strategy in solid tumors. In vivo cell tracking techniques can help us better understand the infiltration, persistence and therapeutic efficacy of CAR T cells. In this field, magnetic resonance imaging (MRI) can achieve high-resolution images of cells by using cellular imaging probes. MRI can also provide various biological information on solid tumors. METHODS: The authors adopted the amino alcohol derivatives of glucose-coated nanoparticles, ultra-small superparamagnetic particles of iron oxide (USPIOs), to label CAR T cells for non-invasive monitoring of kinetic infiltration and persistence in glioblastoma (GBM). The specific targeting CARs included anti-human epidermal growth factor receptor variant III and IL13 receptor subunit alpha 2 CARs. RESULTS: When using an appropriate concentration, USPIO labeling exerted no negative effects on the biological characteristics and killing efficiency of CAR T cells. Increasing hypointensity signals could be detected in GBM models by susceptibility-weighted imaging MRI ranging from 3 days to 14 days following the injection of USPIO-labeled CAR T cells. In addition, nanoparticles and CAR T cells were found on consecutive histopathological sections. Moreover, diffusion and perfusion MRI revealed significantly increased water diffusion and decreased vascular permeability on day 3 after treatment, which was simultaneously accompanied by a significant decrease in tumor cell proliferation and increase in intercellular tight junction on immunostaining sections. CONCLUSION: These results establish an effective imaging technique that can track CAR T cells in GBM models and validate their early therapeutic effects, which may guide the evaluation of CAR T-cell therapies in solid tumors.

10.
Food Chem ; 339: 128151, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33152896

RESUMO

The effects of aloe vera (Aloe vera (L.) Burm. f.) gel treatment on the incidence of superficial scald in 'Starking' apples (Malus domestica Borkh. Var. Starking) during cold storage were studied. Apples were harvested at the pre-climacteric stage and treated with aloe vera gel. The treatment increased malondialdehyde content and membrane lipid damage. Furthermore, it inhibited the release of ethylene at the early stage but increased it in the later stage. The expression level of ACC synthase 1 (MdACS1) also increased, and the antioxidant capacity in apples, particularly, catalase, peroxidase, and superoxide dismutase activities, all decreased, while concomitantly, the content of α-farnesene and its oxidation product, conjugated triene increased, thereby aggravating superficial scald incidence during storage at low temperature.


Assuntos
Conservação de Alimentos/métodos , Malus/fisiologia , Preparações de Plantas , Antioxidantes/metabolismo , Temperatura Baixa , Ciclopropanos/farmacologia , Enzimas/metabolismo , Etilenos/metabolismo , Armazenamento de Alimentos , Frutas/fisiologia , Malus/efeitos dos fármacos , Oxirredução , Preparações de Plantas/farmacologia , Proteínas de Plantas/metabolismo , Sesquiterpenos/metabolismo
11.
IEEE Trans Cybern ; PP2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33259319

RESUMO

This article studies a cooperative output-feedback secure control problem for distributed cyber-physical systems over an unreliable communication interaction, which is to achieve coordination tracking in the presence of intermittent denial-of-service (DoS) attacks. Under the switching communication network environment, first, a distributed secure control method for each subsystem is proposed via neighborhood information, which includes the local state estimator and cooperative resilient controller. Second, based on the topology-dependent Lyapunov function approach, the design conditions of secure control protocol are derived such that cooperative tracking errors are uniformly ultimately bounded. Interestingly, by exploiting the topology-allocation-dependent average dwell-time (TADADT) technique, the stability analysis of closed-loop error dynamics is presented, and the proposed coordination design conditions can relax time constraints on interaction topology switching. Finally, two numerical examples are presented to demonstrate the effectiveness of the theoretical results.

12.
J Orthop Surg Res ; 15(1): 577, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33261646

RESUMO

BACKGROUND: Dexmedetomidine has shown potential in pain control in patients undergoing total knee arthroplasty (TKA). However, the combination of nerve block and dexmedetomidine may be a preferred alternative for postoperative analgesia after TKA. The aim of this study was to perform a meta-analysis on existing randomized controlled trials (RCTs) to determine the efficacy and safety of dexmedetomidine as an adjunct to local anesthetics in nerve block after TKA. METHODS: A literature survey was conducted in the databases of PubMed, Embase, Cochrane Library, Web of science, and ScienceDirect for the RCTs completed before February 1st, 2020 that met pre-specified inclusion criteria. The primary outcomes included the pain scores, duration of analgesia, opioid consumption within 24 h postoperatively, and the level of patient satisfaction. The secondary outcomes included the motor strength, degree of sedation, postoperative nausea and vomiting, and other related complications. The methodological quality was assessed by the Cochrane risk of bias tool. RESULTS: The initial literature search yielded 143 studies, out of which seven studies met the inclusion criteria. The pooled data indicated that dexmedetomidine combined with local anesthetics in nerve block in TKA decreased the postoperative pain scores at rest as well as at motion (SMD = - 1.01 [95% CI - 1.29 to - 0.72], p < 0.01; SMD = - 1.01 [- 1.25 to - 0.77], p < 0.01) respectively, decreased the total opioid consumption within 24 h (SMD = - 0.63 [- 0.86 to - 0.40], p < 0.01), prolonged the duration of analgesia (SMD = 0.90 [0.64 to 1.17], p < 0.01), improved motor strength (SMD = 0.23 [0.01 to 0.45], p = 0.04), improved the degree of sedation (SMD = 0.94 [0.70 to 1.18], p < 0.01), and increased the level of patient satisfaction (SMD = 0.88 [0.60 to 1.17], p < 0.01) without increasing nausea and vomiting (RD = - 0.05 [- 0.11 to 0.01], p = 0.14), as well as other complications (RD = - 0.01 [- 0.08 to 0.07], p = 0.89), compared with local anesthetics alone. CONCLUSIONS: It is effective and safe for dexmedetomidine as an adjunct to local anesthetics in nerve block in TKA to relieve postoperative pain, decrease total opioid consumption, prolong analgesic duration, and increase patient satisfaction without increasing related complications. Based on the quality of evidence, this meta-analysis recommends that dexmedetomidine can be used in a regular treatment regimen and as an adjunct addition to local anesthetics in nerve block for patients undergoing TKA. REGISTRATION: This meta-analysis was prospectively registered on PROSPERO (International prospective register of systematic reviews) and the registering number was CRD42020169171.

14.
Hua Xi Kou Qiang Yi Xue Za Zhi ; 38(6): 681-686, 2020 Dec 01.
Artigo em Chinês | MEDLINE | ID: mdl-33377347

RESUMO

Oral lichen planus (OLP) is a common chronic inflammatory disease of the oral mucosa. The prevalence rate of OLP in adults is 0.5%-2%. The etiology and pathogenesis of OLP are still unclear. The pathogenesis of OLP may be related to the genetic polymorphism of some genes. Currently, the gene families, including tumor necrosis factor, interferon, interleukin, enzyme, and receptor, have been extensively studied. This work reviews related studies on gene polymorphism of OLP.


Assuntos
Líquen Plano Bucal , Adulto , Humanos , Líquen Plano Bucal/genética , Mucosa Bucal , Polimorfismo Genético , Fator de Necrose Tumoral alfa/genética
15.
Commun Biol ; 3(1): 786, 2020 Dec 18.
Artigo em Inglês | MEDLINE | ID: mdl-33339958

RESUMO

A current challenge to produce effective therapeutics is to accurately determine the location of the ligand-biding site and to characterize its properties. So far, the mechanisms underlying the functional activation of cell surface receptors by ligands with a complex binding mechanism remain poorly understood due to a lack of suitable nanoscopic methods to study them in their native environment. Here, we elucidated the ligand-binding mechanism of the human G protein-coupled C5a receptor (C5aR). We discovered for the first time a cooperativity between the two orthosteric binding sites. We found that the N-terminus C5aR serves as a kinetic trap, while the transmembrane domain acts as the functional site and both contributes to the overall high-affinity interaction. In particular, Asp282 plays a key role in ligand binding thermodynamics, as revealed by atomic force microscopy and steered molecular dynamics simulation. Our findings provide a new structural basis for the functional and mechanistic understanding of the GPCR family that binds large macromolecular ligands.

16.
Front Neurol ; 11: 551157, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33224083

RESUMO

Background: Ischemic stroke has a poor prognosis and brings a ponderous burden on families and society. Hemorrhagic transformation (HT) after intravenous thrombolysis can increase the mortality of patients with ischemic stroke. Thus, finding new HT biomarkers to be applicable in clinical practice is of great importance. Methods: The related risk factors were recruited for analysis, including smoking, drinking, hyperlipidemia, diabetes, anamnesis, and pathological indicators. Moreover, the relationship between serum levels of caveolin-1, caveolin-2, and HT after rt-PA treatment were also studied. Results: We studied 306 patients with acute ischemic stroke treated with recombinant tissue type plasminogen activator (rt-PA) within 4.5 h of symptom onset. The results showed that Age ≥68 years, smoking, Atrial fibrillation, NIHSS score before thrombolysis ≥17, and systolic pressure 2 h after thrombolysis (mmHg) ≥149 increased the risks of HT after rt-PA administration. Remarkably, the concentration of caveolin-1 (ng/mL) ≤ 0.12 and caveolin-2 (ng/mL) ≤ 0.43 in serum increased the risks of HT after rt-PA administration. Conclusion: Knowledge on the risk factors associated with HT after rt-PA treatment may help develop treatment strategies and reduce the risk of HT. Caveolin-1 and caveolin-2 can be predictors of HT after rt-PA administration. These findings provide evidence for future further investigations aimed to validate these biomarkers.

17.
Prep Biochem Biotechnol ; : 1-8, 2020 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-33226883

RESUMO

Azo dyes constitute a significant environmental burden due to its toxicity, carcinogenicity, and hard biodegradation. The report here is focused on the decolorization and degradation treatment of azo dye methyl red (MR). Decolorization of MR using Aspergillus versicolor LH1 isolated from activated sludge was investigated. The maximum decolorization rate of 92.3% was obtained under the optimized conditions of sucrose as carbon source, 5d incubation age, pH 6.0, 140 mg/L initial concentration of MR and 2.5 g/L initial concentration of NaNO3. Biodegradation products of MR were investigated using HPLC-MS, FTIR, and GC-MS assays. It was revealed the three bonds of -C-N = in MR aromatic nucleus were disrupted, and benzoic acid was detected. Micronucleus test with Glycine max L. and Vicia faba L. demonstrated that MCN‰ (micronucleus permillage) of MR metabolites was less than MR solution. These findings provided evidence that A. versicolor LH1 is a candidate for MR degradation in industrial wastewater treatment.

18.
Spectrochim Acta A Mol Biomol Spectrosc ; 247: 119105, 2020 Oct 24.
Artigo em Inglês | MEDLINE | ID: mdl-33161265

RESUMO

Hydrogen polysulfides (H2Sn, n ≥ 2) is recently regarded as a potential signaling molecule which shows a higher efficiency than hydrogen sulfides (H2S) in regulating enzymes and ion channels. However, the development of specific fluorescent probes for H2Sn with long-wavelength emission (>600 nm) are still rare. In this work, a semi-naphthorhodafluor-based red-emitting fluorescent probe SNARF-H2Sn containing a phenyl 2-(benzoylthio) benzoate responsive unit was constructed. SNARF-H2Sn was capable of selectively detecting H2Sn over other reactive sulfur species. Treatment with H2Sn would result in a > 1000-fold fluorescence enhancement within 10 min. SNARF-H2Sn showed a low limit of detection down to 6.7 nM, and further enabled to visualize exogenous/endogenous H2Sn in living A549 cells and zebrafish.

19.
Proc Natl Acad Sci U S A ; 117(48): 30755-30762, 2020 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-33199589

RESUMO

Agonist binding promotes activation of G protein-coupled receptors (GPCRs) and association of active receptors with G protein heterotrimers. The resulting active-state ternary complex is the basis for conventional stimulus-response coupling. Although GPCRs can also associate with G proteins before agonist binding, the impact of such preassociated complexes on agonist-induced signaling is poorly understood. Here we show that preassociation of 5-HT7 serotonin receptors with Gs heterotrimers is necessary for agonist-induced signaling. 5-HT7 receptors in their inactive state associate with Gs, as these complexes are stabilized by inverse agonists and receptor mutations that favor the inactive state. Inactive-state 5-HT7-Gs complexes dissociate in response to agonists, allowing the formation of conventional agonist-5-HT7-Gs ternary complexes and subsequent Gs activation. Inactive-state 5-HT7-Gs complexes are required for the full dynamic range of agonist-induced signaling, as 5-HT7 receptors spontaneously activate Gs variants that cannot form inactive-state complexes. Therefore, agonist-induced signaling in this system involves two distinct receptor-G protein complexes, a conventional ternary complex that activates G proteins and an inverse-coupled binary complex that maintains the inactive state when agonist is not present.

20.
Science ; 2020 Nov 05.
Artigo em Inglês | MEDLINE | ID: mdl-33154108

RESUMO

Cost-effective, efficacious therapeutics are urgently needed against the COVID-19 pandemic. Here, we used camelid immunization and proteomics to identify a large repertoire of highly potent neutralizing nanobodies (Nbs) to the SARS-CoV-2 spike (S) protein receptor-binding domain (RBD). We discovered Nbs with picomolar to femtomolar affinities that inhibit viral infection at sub-ng/ml concentration and determined a structure of one of the most potent in complex with RBD. Structural proteomics and integrative modeling revealed multiple distinct and non-overlapping epitopes and indicated an array of potential neutralization mechanisms. We constructed multivalent Nb constructs that achieved ultrahigh neutralization potency (IC50s as low as 0.058 ng/ml) and may prevent mutational escape. These thermostable Nbs can be rapidly produced in bulk from microbes and resist lyophilization, and aerosolization.

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