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APS1/APECED patients are defined by defects in the autoimmune regulator (AIRE) that mediates central T cell tolerance to many self-antigens. AIRE deficiency also affects B cell tolerance, but this is incompletely understood. Here we show that most APS1/APECED patients displayed B cell autoreactivity toward unique sets of approximately 100 self-proteins. Thereby, autoantibodies from 81 patients collectively detected many thousands of human proteins. The loss of B cell tolerance seemingly occurred during antibody affinity maturation, an obligatorily T cell-dependent step. Consistent with this, many APS1/APECED patients harbored extremely high-affinity, neutralizing autoantibodies, particularly against specific cytokines. Such antibodies were biologically active in vitro and in vivo, and those neutralizing type I interferons (IFNs) showed a striking inverse correlation with type I diabetes, not shown by other anti-cytokine antibodies. Thus, naturally occurring human autoantibodies may actively limit disease and be of therapeutic utility.
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Afinidad de Anticuerpos , Autoanticuerpos/inmunología , Resistencia a la Enfermedad/inmunología , Poliendocrinopatías Autoinmunes/inmunología , Factores de Transcripción/deficiencia , Adolescente , Adulto , Anciano , Animales , Anticuerpos Neutralizantes/inmunología , Niño , Preescolar , Citocinas/inmunología , Diabetes Mellitus Tipo 1/inmunología , Humanos , Tolerancia Inmunológica , Ratones Endogámicos C57BL , Persona de Mediana Edad , Linfocitos T/inmunología , Adulto Joven , Proteína AIRERESUMEN
RNA vaccines elicit protective immunity against SARS-CoV-2, but the use of mRNA as an antiviral immunotherapeutic is unexplored. Here, we investigate the activity of lipidoid nanoparticle (LNP)-formulated mRNA encoding human IFNλ1 (ETH47), which is a critical driver of innate immunity at mucosal surfaces protecting from viral infections. IFNλ1 mRNA administration promotes dose-dependent protein translation, induction of interferon-stimulated genes without relevant signs of unspecific immune stimulation, and dose-dependent inhibition of SARS-CoV-2 replication in vitro. Pulmonary administration of IFNλ1 mRNA in mice results in a potent reduction of virus load, virus-induced body weight loss and significantly increased survival. These data support the development of inhaled administration of IFNλ1 mRNA as a potential prophylactic option for individuals exposed to SARS-CoV-2 or at risk suffering from COVID-19. Based on the broad antiviral activity of IFNλ1 regardless of virus or variant, this approach might also be utilized for other respiratory viral infections or pandemic preparedness.
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This corrects the article DOI: 10.1103/PhysRevLett.125.123003.
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We perform high-resolution spectroscopy of the 3d ^{2}D_{3/2}-3d ^{2}D_{5/2} interval in all stable even isotopes of ^{A}Ca^{+} (A=40, 42, 44, 46, and 48) with an accuracy of â¼20 Hz using direct frequency-comb Raman spectroscopy. Combining these data with isotope shift measurements of the 4s ^{2}S_{1/2}â3d ^{2}D_{5/2} transition, we carry out a King plot analysis with unprecedented sensitivity to coupling between electrons and neutrons by bosons beyond the standard model. Furthermore, we estimate the sensitivity to such bosons from equivalent spectroscopy in Ba^{+} and Yb^{+}. Finally, the data yield isotope shifts of the 4s ^{2}S_{1/2}â3d ^{2}D_{3/2} transition at 10 parts per billion through combination with recent data of Knollmann, Patel, and Doret [Phys. Rev. A 100, 022514 (2019)PLRAAN2469-992610.1103/PhysRevA.100.022514].
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In biology, high valent oxo-iron(IV) species have been shown to be pivotal intermediates for functionalization of C-H bonds in the catalytic cycles of a range of O2-activating iron enzymes. This work details an electronic-structure investigation of [FeIV(O)(LNHC)(NCMe)]2+ (LNHC = 3,9,14,20-tetraaza-1,6,12,17-tetraazoniapenta-cyclohexacosane-1(23),4,6(26),10,12(25),15,17(24),21-octaene, complex 1) using helium tagging infrared photodissociation (IRPD), absorption, and magnetic circular dichroism (MCD) spectroscopy, coupled with DFT and highly correlated wave function based multireference calculations. The IRPD spectrum of complex 1 reveals the Fe-O stretching vibration at 832 ± 3 cm-1. By analyzing the Franck-Condon progression, we can determine the same vibration occurring at 616 ± 10 cm-1 in the E(dxy â dxz,yz) excited state. Both values are similar to those measured for [FeIV(O)(TMC)(NCMe)]2+ (TMC = 1,4,8,11-tetramethyl-1,4,8,11-tetraazacyclotetradecane). The low-temperature MCD spectra of complex 1 exhibit three pseudo A-term signals around 12â¯500, 17â¯000, and 24â¯300 cm-1. We can unequivocally assign them to the ligand field transitions of dxy â dxz,yz, dxz,yz â dz2, and dxz,yz â dx2-y2, respectively, through direct calculations of MCD spectra and independent determination of the MCD C-term signs from the corresponding electron donating and accepting orbitals. In comparison with the corresponding transitions observed for [FeIV(O) (SR-TPA)(NCMe)]2+ (SR-TPA = tris(3,5-dimethyl-4-methoxypyridyl-2-methy)amine), the excitations within the (FeO)2+ core of complex 1 have similar transition energies, whereas the excitation energy for dxz,yz â dx2-y2 is significantly higher (â¼12â¯000 cm-1 for [FeIV(O)(SR-TPA)(NCMe)]2+). Our results thus substantiate that the tetracarbene ligand (LNHC) of complex 1 does not significantly affect the bonding in the (FeO)2+ unit but strongly destabilizes the dx2-y2 orbital to eventually lift it above dz2. As a consequence, this unusual electron configuration leads to an unprecedentedly larger quintet-triplet energy separation for complex 1, which largely rules out the possibility that the H atom transfer reaction may take place on the quintet surface and hence quenches two-state reactivity. The resulting mechanistic implications are discussed.
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Amidines have found widespread use, but their solution chemistry remains poorly understood. In this work, X-ray crystallographic and detailed 1D and 2D NMR spectroscopic studies have been performed to elucidate the preferred isomers and their interconversion mechanisms. Amidines are shown to exist as a mixture of E-syn and Z-anti isomers in solution and to form dimeric H-bonded aggregates that are also observed in the solid state. Rapid proton exchange/tautomerization reactions occur within the dimers, allowing fast interconversion of E-syn and Z-anti isomers even at very low temperatures. Three different exchange processes were identified in solution, and on this basis the unusual concentration and temperature dependence of the NMR spectra of these amidines could be explained. This work thus resolves some of the puzzles of the complex solution chemistry of this prominent class of compounds.
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A disulfide-bridged diiron complex with [Fe-S-S-Fe] core, which represents an isomer of the common biological [2Fe-2S] ferredoxin-type clusters, was synthesized using strongly σ-donating macrocyclic tetracarbene capping ligands. Though the complex is quite labile in solution, single crystals were obtained, and the structure was elucidated by X-ray diffraction. The electron-rich iron-sulfur core is found to show rather unusual magnetic and electronic properties. Experimental data and density functional theory studies indicate extremely strong antiferromagnetic coupling (-J > 800 cm(-1)) between two low-spin iron(III) ions via the S2(2-) bridge, and the intense near-IR absorption characteristic for the [Fe-S-S-Fe] core was assigned to a S â Fe ligand-to-metal charge transfer transition.
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Disulfuros/química , Compuestos Férricos/química , Cristalografía por Rayos X , Dimerización , Electrones , Compuestos Férricos/síntesis química , Modelos Moleculares , Estructura Molecular , Teoría CuánticaRESUMEN
A new powerful and oxidatively rugged pyrazolate-based water oxidation catalyst of formula {[Ru(II)(py-SO3)2(H2O)]2(µ-Mebbp)}(-), 1(H2O)2(-), has been prepared and thoroughly characterized spectroscopically and electrochemically. This new catalyst has been conceived based on a specific ligand tailoring design, so that its performance has been systematically improved. It was also demonstrated how subtle ligand modifications cause a change in the O-O bond formation mechanism, thus revealing the close activation energy barriers associated with each pathway.
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A new pyrazole ligand with flexible thioether chelate arms was synthesized and was used to obtain an unprecedented class of hexanuclear coinage metal complexes of general formula [MM'L]3Y3 (M, M' = Cu, Ag, Au; Y = OTf, BF4). Three of them were characterized by X-ray crystallography, namely, homometallic [Ag2L]3(OTf)3 and [Ag2L]3(BF4)3 as well as heterometallic [CuAgL]3(OTf)3, revealing that the classical [M(µ-pz)]3 core is crowned by a second deck of S-bound M' ions. Depending on the solvent, these oligonuclear systems undergo rapid dynamics and show cation-anion aggregation in solution, which has been investigated by DOSY and temperature dependent NMR spectroscopy. Preliminary luminescence data for selected hexametallic [MM'L]3Y3 complexes show that the combination of ligand-directed intramolecular and supramolecular d(10) metal ion interactions in the solid state gives rise to synergic emissive properties that allow for a selective addressing of different emission wavelengths.
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1,3-Bis(pentafluorophenyl-imino)isoindoline (A(F)) and 3,6-di-tert-butyl-1,8-bis(pentafluorophenyl)-9H-carbazole (B(F)) have been designed as preorganized anion receptors that exploit anion-π interactions, and their ability to bind chloride and bromide in various solvents has been evaluated. Both receptors A(F) and B(F) are neutral but provide a central NH hydrogen bond that directs the halide anion into a preorganized clamp of the two electron-deficient appended arenes. Crystal structures of host-guest complexes of A(F) with DMSO, Cl(-), or Br(-) (A(F):DMSO, A(F):Cl(-), and A(2)(F):Br(-)) reveal that in all cases the guest is located in the cleft between the perfluorinated flaps, but NMR spectroscopy shows a more complex situation in solution because of E,Z/Z,Z isomerism of the host. In the case of the more rigid receptor B(F), Job plots evidence 1:1 complex formation with Cl(-) and Br(-), and association constants up to 960 M(-1) have been determined depending on the solvent. Crystal structures of B(F) and B(F):DMSO visualize the distinct preorganization of the host for anion-π interactions. The reference compounds 1,3-bis(2-pyrimidylimino)isoindoline (A(N)) and 3,6-di-tert-butyl-1,8-diphenyl-9H-carbazole (B(H)), which lack the perfluorinated flaps, do not show any indication of anion binding under the same conditions. A detailed computational analysis of the receptors A(F) and B(F) and their host-guest complexes with Cl(-) or Br(-) was carried out to quantify the interactions in play. Local correlation methods were applied, allowing for a decomposition of the ring-anion interactions. The latter were found to contribute significantly to the stabilization of these complexes (about half of the total energy). Compounds A(F) and B(F) represent rare examples of neutral receptors that are well preorganized for exploiting anion-π interactions, and rare examples of receptors for which the individual contributions to the binding energy have been quantified.
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A new family of tetranuclear nickel cube complexes [Ni(4)L(4)(solv)(4)] (1, solv = MeOH; 2, solv = H(2)O; H(2)L = pyrazole-based tridentate {ONO} ligand) has been studied in detail, in particular by X-ray diffraction and superconducting quantum interference device (SQUID) magnetometry. Different solvates 1·H(2)O, 2·4C(3)H(6)O, 2·CH(2)Cl(2), and 2·H(2)O were obtained in crystalline form. Only small structural variations were found for the Ni-O-Ni angles of the [Ni(4)O(4)] cores of those compounds, but these slight variations have dramatic consequences for the magnetic properties. [Ni(4)L(4)(MeOH)(4)]·H(2)O (1·H(2)O) and [Ni(4)L(4)(H(2)O)(4)]·H(2)O (2·H(2)O) can be reversibly interconverted in the solid state by exposure to the respective solvent, MeOH or H(2)O, and this goes along with a switching of the spin ground state from magnetic (S(T) = 4) to diamagnetic (S(T) = 0). Likewise the (irreversible) loss of lattice solvent in [Ni(4)L(4)(H(2)O)(4)]·4C(3)H(6)O (2·4C(3)H(6)O) to give 2·2C(3)H(6)O changes the ground state from S(T) = 4 to S(T) = 0. In view of these dramatic solvatomagnetic effects for the present [Ni(4)L(4)(solv)(4)] complexes, which occur upon extrusion of lattice solvent or facile exchange of coordinated solvent molecules while keeping the robust [Ni(4)O(4)] core intact, a note of care is issued: whenever magnetic data are obtained for powdered material or for crystals that easily loose lattice solvent molecules, the magnetic properties may not necessarily reflect the situation observed in the corresponding single crystal diffraction study. Finally, a thorough analysis of the present series of complexes as well as other {Ni(4)(µ(3)-OR)(4)} cubes reported in the literature confirms that a correlation between the (Ni-O-Ni)(av) bond angle and J in [Ni(4)O(4)] cubane complexes does indeed exist.
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Complejos de Coordinación/síntesis química , Hierro/química , Metano/análogos & derivados , Complejos de Coordinación/química , Cristalografía por Rayos X , Metano/química , Conformación Molecular , Compuestos Organometálicos/síntesis química , Compuestos Organometálicos/química , Espectroscopía de MossbauerRESUMEN
A sucrose-inducible promoter system (P(sacB)) from Bacillus megaterium was identified using a secretome approach. It was successfully employed for the extracellular production of the homologous levansucrase SacB (4252.4 U l(-1)) and the heterologous green fluorescent protein GFP (7.9 mg g(CDW)(-1)). Mutational analysis of B. megaterium P(sacB) allowed the identification of important promoter elements. The sucrose-inducible promoter provides a useful alternative to the established xylose-inducible promoter system (P(xylA)) for recombinant gene expression in B. megaterium.
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Bacillus megaterium/metabolismo , Proteínas Fluorescentes Verdes/metabolismo , Hexosiltransferasas/metabolismo , Proteínas Recombinantes/metabolismo , Sacarosa/metabolismo , Análisis Mutacional de ADN , Regiones Promotoras GenéticasRESUMEN
Device scale-up and long-term stability constitute two major hurdles that the emerging perovskite solar technology will have to overcome before commercialization. Here, a comparative study was performed between ZnO and TiO2 electron-selective layers, two materials that allow the low-temperature processing of perovskite solar cells on polymer substrates. Although the use of TiO2 is well established on glass substrates, ZnO was chosen because it can be readily printed at low temperature and offers the potential for the large-scale roll-to-roll manufacturing of flexible photovoltaics at a low cost. However, a rapid degradation of CH3 NH3 PbI3 was observed if it was deposited on ZnO, therefore, the influence of the perovskite film preparation conditions on its morphology and degradation kinetics was investigated. This study showed that CH3 NH3 PbI3 could withstand a higher temperature on TiO2 than ZnO and that TiO2-based perovskite devices were more stable than their ZnO analogues.
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Compuestos de Calcio/química , Suministros de Energía Eléctrica , Óxidos/química , Polímeros/química , Energía Solar , Titanio/química , Óxido de Zinc/química , Microscopía Electrónica de Rastreo , Difracción de Rayos XRESUMEN
The Baculovirus Expression Vector System (BEVS) is widely used to produce high amounts of recombinant proteins. Nevertheless, generating recombinant baculovirus in high quality is rather time-consuming and labor-intensive. Alternatively, virus-free expression in insect cells did not achieve similar expression levels for most proteins so far. The transactivation method is a promising approach for protein expression in Sf21 cells. It combines advantages of BEVS and plasmid-based expression by activating strong virus-dependent promoters on a transfected plasmid by baculoviral coinfection. Here, we identified expression elements required for transactivation. Therefore, we designed several vectors comprising different viral promoters or promoter combinations and tested them for eGFP expression using the automated BioLector microcultivation system. Remarkably, only the combination of the very late promoter p10 together with the homologous region 5 (hr5) could boost expression during transactivation. Other elements, like p10 alone or the late viral promoter polH, did not respond to transactivation. A new combination of hr5 and p10 with the strongest immediate early OpMNPV viral promoter OpIE2 improved the yield of eGFP by ~25% in comparison to the previous applied hr5-IE1-p10 expression cassette. Furthermore, we observed a strong influence of the transcription termination sequence and vector backbone on the level of expression. Finally, the expression levels for transactivation, BEVS and solely plasmid-based expression were compared for the marker protein eGFP, underlining the potential of transactivation for fast recombinant protein expression in Sf21 cells. In conclusion, essential elements for transactivation could be identified. The optimal elements were applied to generate an improved vector applicable in virus-free plasmid-based expression, transactivation and BEVS.
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Baculoviridae/genética , Insectos/genética , Proteínas Recombinantes/genética , Activación Transcripcional/genética , Animales , Línea Celular , Genes Reporteros/genética , Vectores Genéticos/genética , Proteínas Fluorescentes Verdes/genética , Plásmidos/genética , Regiones Promotoras Genéticas/genética , Células Sf9 , Transcripción Genética/genética , Transfección/métodosRESUMEN
The objective was to assess the long-term outcome of secondary alveolar bone grafting (SABG) in cleft lip and palate patients and to examine relationships between preoperative and postoperative factors and overall long-term bone graft success. The records of 97 patients with cleft lip and palate, who had secondary alveolar bone grafting of 123 alveolar clefts, were examined. Interalveolar bone height was assessed radiographically a minimum of 10 years after grafting using a 4-point scale (I-IV), where types I and II were considered a success. After an average follow-up of 16 years after SABG (range = 10.2-22.7 years), 101 of the 123 grafts (82%) were categorised as successes. Mean age in the success group was 12.1 years and 13.6 years in the failure group (p = 0.03). It was found that the success rate was significantly lower (p = 0.02) if SABG was performed after eruption of the tooth distal to the cleft. No significant differences were found with regard to the other parameters investigated. The timing of secondary alveolar bone grafting is critical with regard to the age of the patient and the stage of eruption of the tooth distal to the cleft.
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Injerto de Hueso Alveolar , Labio Leporino/cirugía , Fisura del Paladar/cirugía , Evaluación del Resultado de la Atención al Paciente , Adolescente , Factores de Edad , Proceso Alveolar/diagnóstico por imagen , Niño , Estudios de Cohortes , Femenino , Estudios de Seguimiento , Humanos , Masculino , Ortodoncia Correctiva , Radiografía , Factores Sexuales , Erupción DentalRESUMEN
Recombinant production of complex eukaryotic proteins for structural analyses typically requires a profound screening process to identify suitable constructs for the expression of ample amounts of properly folded protein. Furthermore, the evaluation of an optimal expression host has a major impact on protein yield and quality as well as on actual cost of the production process. Here we present a novel fast expression system for multiple hosts based on a single donor vector termed pFlp-Bac-to-Mam. The range of applications of pFlp-Bac-to-Mam comprises highly efficient transient transfection of HEK293-6E in serum-free suspension culture and subsequent large-scale production of challenging proteins expressing in mg per Liter level using either the baculoviral expression vector system or stable CHO production cell lines generated by Flp-mediated cassette exchange. The success of the multi-host expression vector to identify the optimal expression strategy for efficient production of high quality protein is demonstrated in a comparative expression study of three model proteins representing different protein classes: intracellular expression using a fluorescent protein, secretion of a single-chain-Fv-hIgG1Fc fusion construct and production of a large amount of highly homogeneous protein sample of the extracellular domain of a Toll-like receptor. The evaluation of the production efficiency shows that the pFlp-Bac-to-Mam system allows a fast and individual optimization of the expression strategy for each protein class.