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1.
Int Immunopharmacol ; 129: 111569, 2024 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-38340419

RESUMO

The COVID-19 pandemic has underscored the pressing need for safe and effective booster vaccines, particularly in considering the emergence of new SARS-CoV-2 variants and addressing vaccine distribution inequalities. Dissolving microneedle array patches (MAP) offer a promising delivery method, enhancing immunogenicity and improving accessibility through the skin's immune potential. In this study, we evaluated a microneedle array patch-based S1 subunit protein COVID-19 vaccine candidate, which comprised a bivalent formulation targeting the Wuhan and Beta variant alongside a monovalent Delta variant spike proteins in a murine model. Notably, the second boost of homologous bivalent MAP-S1(WU + Beta) induced a 15.7-fold increase in IgG endpoint titer, while the third boost of heterologous MAP-S1RS09Delta yielded a more modest 1.6-fold increase. Importantly, this study demonstrated that the administration of four doses of the MAP vaccine induced robust and long-lasting immune responses, persisting for at least 80 weeks. These immune responses encompassed various IgG isotypes and remained statistically significant for one year. Furthermore, neutralizing antibodies against multiple SARS-CoV-2 variants were generated, with comparable responses observed against the Omicron variant. Overall, these findings emphasize the potential of MAP-based vaccines as a promising strategy to combat the evolving landscape of COVID-19 and to deliver a safe and effective booster vaccine worldwide.


Assuntos
Vacinas contra COVID-19 , COVID-19 , Animais , Humanos , Camundongos , Subunidades Proteicas , SARS-CoV-2 , Pandemias , COVID-19/prevenção & controle , Anticorpos Neutralizantes , Imunoglobulina G , Anticorpos Antivirais
2.
Food Chem ; 441: 138371, 2024 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-38218148

RESUMO

The qualities of wheat dough are influenced by the high-molecular-weight glutenin subunits (HMW-GS), a critical component of wheat gluten protein. However, it is still unknown how HMW-GS silencing affects the aggregation characteristics of dough. Two groups of near-isogenic wheat were used to study the effects of HMW-GS silencing on dough aggregation characteristics, dough texture characteristics, and dough microstructure. It was observed that the content of gliadin in LH-11 strain significantly increased compared to the wild-type (WT). Additionally, the amount of glutenin macropolymer and the glutenin/gliadin both decreased. The aggregation characteristics and rheological characteristics of the dough in LH-11 strain were significantly reduced, and the content of ß-sheet in the dough was significantly reduced. The HMW-GS silencing resulted in a reduction in the aggregation of the gluten network in the dough, which related to the alteration of the secondary and microstructure of the gluten.


Assuntos
Gliadina , Glutens , Gliadina/metabolismo , Peso Molecular , Glutens/química , Triticum/química , Farinha , Subunidades Proteicas/química
3.
J Am Chem Soc ; 146(6): 3984-3991, 2024 Feb 14.
Artigo em Inglês | MEDLINE | ID: mdl-38236721

RESUMO

The light-harvesting antennae of diatoms and spinach are composed of similar chromophores; however, they exhibit different absorption wavelengths. Recent advances in cryoelectron microscopy have revealed that the diatom light-harvesting antenna fucoxanthin chlorophyll a/c-binding protein (FCPII) forms a tetramer and differs from the spinach antenna in terms of the number of protomers; however, the detailed molecular mechanism remains elusive. Herein, we report the physicochemical factors contributing to the characteristic light absorption of the diatom light-harvesting antenna based on spectral calculations using an exciton model. Spectral analysis reveals the significant contribution of unique fucoxanthin molecules (fucoxanthin-S) in FCPII to the diatom-specific spectrum, and further analysis determines their essential role in excitation-energy transfer to chlorophyll. It was revealed that the specificity of these fucoxanthin-S molecules is caused by the proximity between protomers associated with the tetramerization of FCPII. The findings of this study demonstrate that diatoms employ fucoxanthin-S to harvest energy under the ocean in the absence of long-wavelength sunlight and can provide significant information about the survival strategies of photosynthetic organisms to adjust to their living environment.


Assuntos
Carotenoides , Diatomáceas , Xantofilas , Carotenoides/química , Clorofila A , Diatomáceas/química , Microscopia Crioeletrônica , Subunidades Proteicas/metabolismo , Clorofila/química , Complexos de Proteínas Captadores de Luz/química , Transferência de Energia , Proteínas de Ligação à Clorofila/química , Proteínas de Ligação à Clorofila/metabolismo
4.
J Biol Chem ; 300(1): 105576, 2024 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-38110033

RESUMO

The sixth family phosphodiesterases (PDE6) are principal effector enzymes of the phototransduction cascade in rods and cones. Maturation of nascent PDE6 protein into a functional enzyme relies on a coordinated action of ubiquitous chaperone HSP90, its specialized cochaperone aryl hydrocarbon receptor-interacting protein-like 1 (AIPL1), and the regulatory Pγ-subunit of PDE6. Deficits in PDE6 maturation and function underlie severe visual disorders and blindness. Here, to elucidate the roles of HSP90, AIPL1, and Pγ in the maturation process, we developed the heterologous expression system of human cone PDE6C in insect cells allowing characterization of the purified enzyme. We demonstrate that in the absence of Pγ, HSP90, and AIPL1 convert the inactive and aggregating PDE6C species into dimeric PDE6C that is predominantly misassembled. Nonetheless, a small fraction of PDE6C is properly assembled and fully functional. From the analysis of mutant mice that lack both rod Pγ and PDE6C, we conclude that, in contrast to the cone enzyme, no maturation of rod PDE6AB occurs in the absence of Pγ. Co-expression of PDE6C with AIPL1 and Pγ in insect cells leads to a fully mature enzyme that is equivalent to retinal PDE6. Lastly, using immature PDE6C and purified chaperone components, we reconstituted the process of the client maturation in vitro. Based on this analysis we propose a scheme for the PDE6 maturation process.


Assuntos
Nucleotídeo Cíclico Fosfodiesterase do Tipo 6 , Células Fotorreceptoras Retinianas Cones , Animais , Humanos , Camundongos , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Cegueira/genética , Linhagem Celular , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/química , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/deficiência , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/genética , Nucleotídeo Cíclico Fosfodiesterase do Tipo 6/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Mutação , Multimerização Proteica , Subunidades Proteicas/química , Subunidades Proteicas/deficiência , Subunidades Proteicas/genética , Subunidades Proteicas/metabolismo , Células Fotorreceptoras Retinianas Cones/química , Células Fotorreceptoras Retinianas Cones/metabolismo
5.
Microb Cell Fact ; 22(1): 244, 2023 Nov 30.
Artigo em Inglês | MEDLINE | ID: mdl-38037098

RESUMO

Protein complex assembly facilitates the combination of individual protein subunits into functional entities, and thus plays a crucial role in biology and biotechnology. Recently, we developed quasi-twodimensional, silicon-based compartmental biochips that are designed to study and administer the synthesis and assembly of protein complexes. At these biochips, individual protein subunits are synthesized from locally confined high-density DNA brushes and are captured on the chip surface by molecular traps. Here, we investigate single-gene versions of our quasi-twodimensional synthesis systems and introduce the trap-binding efficiency to characterize their performance. We show by mathematical and computational modeling how a finite trap density determines the dynamics of protein-trap binding and identify three distinct regimes of the trap-binding efficiency. We systematically study how protein-trap binding is governed by the system's three key parameters, which are the synthesis rate, the diffusion constant and the trap-binding affinity of the expressed protein. In addition, we describe how spatially differential patterns of traps modulate the protein-trap binding dynamics. In this way, we extend the theoretical knowledge base for synthesis, diffusion, and binding in compartmental systems, which helps to achieve better control of directed molecular self-assembly required for the fabrication of nanomachines for synthetic biology applications or nanotechnological purposes.


Assuntos
Nanotecnologia , Biossíntese de Proteínas , Subunidades Proteicas , Nanotecnologia/métodos , Simulação por Computador , Silício
6.
Arq Bras Cardiol ; 120(12): e20230396, 2023 Dec.
Artigo em Português, Inglês | MEDLINE | ID: mdl-38126445

RESUMO

BACKGROUND: Central Illustration : G Protein Subunit Beta 3 (GNB3) Variant Is Associated with Biochemical Changes in Brazilian Patients with Hypertension. BACKGROUND: Genes and their variants associated with environmental factors contribute to the development of the hypertensive phenotype. The G protein beta 3 subunit gene (GNB3) is involved in the intracellular signaling process, and its variants have been related to susceptibility to arterial hypertension. OBJECTIVE: To determine the association of the GNB3 variant (rs5443:C>T) with arterial hypertension, biochemical parameters, age, and obesity in hypertensive and normotensive individuals from Ouro Preto, Minas Gerais, Brazil. METHOD: The identification of variants was performed by real-time PCR, using the TaqMan® system, in 310 samples (155 hypertensive and 155 normotensive). Biochemical analyses (renal function, lipid profile and glycemia) were performed from the serum using UV/Vis spectrophotometry and ion-selective electrode. A multiple logistic regression model was used to identify factors associated with arterial hypertension. The analysis of continuous variables with normal distribution was performed using the unpaired Student's t test; non-normal data were analyzed using Mann-Whitney. P < 0.05 was considered significant. RESULTS: The rs5443:C>T variant was not associated with arterial hypertension in the evaluated population (p = 0.88). Regarding biochemical measures, the T allele was associated with high levels of triglycerides, glucose and uric acid in hypertensive individuals (p < 0.05). CONCLUSION: These results show the importance of genetic diagnosis to prevent the causes and consequences of diseases and imply that the GNB3 rs5443:C>T variant may be associated with changes in the biochemical profile in hypertensive individuals.


Assuntos
Proteínas Heterotriméricas de Ligação ao GTP , Hipertensão , Humanos , Alelos , Pressão Sanguínea/genética , Brasil , Genótipo , Hipertensão/genética , Subunidades Proteicas/genética , Proteínas Heterotriméricas de Ligação ao GTP/genética
7.
Science ; 382(6677): 1404-1411, 2023 12 22.
Artigo em Inglês | MEDLINE | ID: mdl-38127736

RESUMO

Gain-of-function mutations in LRRK2, which encodes the leucine-rich repeat kinase 2 (LRRK2), are the most common genetic cause of late-onset Parkinson's disease. LRRK2 is recruited to membrane organelles and activated by Rab29, a Rab guanosine triphosphatase encoded in the PARK16 locus. We present cryo-electron microscopy structures of Rab29-LRRK2 complexes in three oligomeric states, providing key snapshots during LRRK2 recruitment and activation. Rab29 induces an unexpected tetrameric assembly of LRRK2, formed by two kinase-active central protomers and two kinase-inactive peripheral protomers. The central protomers resemble the active-like state trapped by the type I kinase inhibitor DNL201, a compound that underwent a phase 1 clinical trial. Our work reveals the structural mechanism of LRRK2 spatial regulation and provides insights into LRRK2 inhibitor design for Parkinson's disease treatment.


Assuntos
Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina , Doença de Parkinson , Proteínas rab de Ligação ao GTP , Humanos , Antiparkinsonianos/química , Antiparkinsonianos/farmacologia , Domínio Catalítico , Microscopia Crioeletrônica , Desenho de Fármacos , Mutação com Ganho de Função , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/antagonistas & inibidores , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/química , Serina-Treonina Proteína Quinase-2 com Repetições Ricas em Leucina/genética , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/genética , Subunidades Proteicas/química , Proteínas rab de Ligação ao GTP/química , Multimerização Proteica , Inibidores de Proteínas Quinases/química , Inibidores de Proteínas Quinases/farmacologia
8.
Curr Opin Neurobiol ; 83: 102806, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37950957

RESUMO

N-methyl-d-aspartate receptors (NMDARs) belong to the ionotropic glutamate receptors (iGluRs) superfamily and act as coincidence detectors that are crucial to neuronal development and synaptic plasticity. They typically assemble as heterotetramers of two obligatory GluN1 subunits and two alternative GluN2 (from 2A to 2D) and/or GluN3 (3A and 3B) subunits. These alternative subunits mainly determine the diverse biophysical and pharmacological properties of different NMDAR subtypes. Over the past decade, the unprecedented advances in structure elucidation of these tetrameric NMDARs have provided atomic insights into channel gating, allosteric modulation and the action of therapeutic drugs. A wealth of structural and functional information would accelerate the artificial intelligence-based drug design to exploit more NMDAR subtype-specific molecules for the treatment of neurological and psychiatric disorders.


Assuntos
Inteligência Artificial , Receptores de N-Metil-D-Aspartato , Humanos , Receptores de N-Metil-D-Aspartato/metabolismo , Regulação Alostérica , Subunidades Proteicas/química , Subunidades Proteicas/metabolismo
9.
Cell Chem Biol ; 30(11): 1329-1331, 2023 11 16.
Artigo em Inglês | MEDLINE | ID: mdl-37977126

RESUMO

The precise and selective quantification of drug-target interactions within the context of RAS-RAF heterodimers in live cells offers a powerful tool for drug development and personalized medicine, particularly in cancer research, where the RAS-RAF pathway is pivotal.


Assuntos
Fagocitose , Subunidades Proteicas
10.
Cells ; 12(19)2023 10 07.
Artigo em Inglês | MEDLINE | ID: mdl-37830628

RESUMO

Monomers, dimers, and individual FOF1-ATP synthase subunits are, presumably, involved in the formation of the mitochondrial permeability transition pore (PTP), whose molecular structure, however, is still unknown. We hypothesized that, during the Ca2+-dependent assembly of a PTP complex, the F-ATP synthase (subunits) recruits mitochondrial proteins that do not interact or weakly interact with the F-ATP synthase under normal conditions. Therefore, we examined whether the PTP opening in mitochondria before the separation of supercomplexes via BN-PAGE will increase the channel stability and channel-forming capacity of isolated F-ATP synthase dimers and monomers in planar lipid membranes. Additionally, we studied the specific activity and the protein composition of F-ATP synthase dimers and monomers from rat liver and heart mitochondria before and after PTP opening. Against our expectations, preliminary PTP opening dramatically suppressed the high-conductance channel activity of F-ATP synthase dimers and monomers and decreased their specific "in-gel" activity. The decline in the channel-forming activity correlated with the reduced levels of as few as two proteins in the bands: methylmalonate-semialdehyde dehydrogenase and prohibitin 2. These results indicate that proteins co-migrating with the F-ATP synthase may be important players in PTP formation and stabilization.


Assuntos
Proteínas de Transporte da Membrana Mitocondrial , ATPases Mitocondriais Próton-Translocadoras , ATPases Mitocondriais Próton-Translocadoras/metabolismo , Proteínas de Transporte da Membrana Mitocondrial/metabolismo , Subunidades Proteicas/metabolismo , Mitocôndrias Cardíacas/metabolismo , Trifosfato de Adenosina
11.
J Biol Chem ; 299(11): 105318, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37797699

RESUMO

Collagen IV scaffold is a primordial innovation enabling the assembly of a fundamental architectural unit of epithelial tissues-a basement membrane attached to polarized cells. A family of six α-chains (α1 to α6) coassemble into three distinct protomers that form supramolecular scaffolds, noted as collagen IVα121, collagen IVα345, and collagen IVα121-α556. Chloride ions play a pivotal role in scaffold assembly, based on studies of NC1 hexamers from mammalian tissues. First, Cl- activates a molecular switch within trimeric NC1 domains that initiates protomer oligomerization, forming an NC1 hexamer between adjoining protomers. Second, Cl- stabilizes the hexamer structure. Whether this Cl--dependent mechanism is of fundamental importance in animal evolution is unknown. Here, we developed a simple in vitro method of SDS-PAGE to determine the role of solution Cl- in hexamer stability. Hexamers were characterized from 34 animal species across 15 major phyla, including the basal Cnidarian and Ctenophora phyla. We found that solution Cl- stabilized the quaternary hexamer structure across all phyla except Ctenophora, Ecdysozoa, and Rotifera. Further analysis of hexamers from peroxidasin knockout mice, a model for decreasing hexamer crosslinks, showed that solution Cl- also stabilized the hexamer surface conformation. The presence of sufficient chloride concentration in solution or "chloride pressure" dynamically maintains the native form of the hexamer. Collectively, our findings revealed that chloride pressure on the outside of cells is a primordial innovation that drives and maintains the quaternary and conformational structure of NC1 hexamers of collagen IV scaffolds.


Assuntos
Cloretos , Colágeno Tipo IV , Animais , Camundongos , Subunidades Proteicas/análise , Estrutura Terciária de Proteína , Colágeno Tipo IV/química , Membrana Basal , Mamíferos
12.
PLoS Comput Biol ; 19(10): e1011545, 2023 10.
Artigo em Inglês | MEDLINE | ID: mdl-37831724

RESUMO

TRPV Ion channels are sophisticated molecular sensors designed to respond to distinct temperature thresholds. The recent surge in cryo-EM structures has provided numerous insights into the structural rearrangements accompanying their opening and closing; however, the molecular mechanisms by which TRPV channels establish precise and robust temperature sensing remain elusive. In this work we employ molecular simulations, multi-ensemble contact analysis, graph theory, and machine learning techniques to reveal the temperature-sensitive residue-residue interactions driving allostery in TRPV3. We find that groups of residues exhibiting similar temperature-dependent contact frequency profiles cluster at specific regions of the channel. The dominant mode clusters on the ankyrin repeat domain and displays a linear melting trend while others display non-linear trends. These modes describe the residue-level temperature response patterns that underlie the channel's functional dynamics. With network analysis, we find that the community structure of the channel changes with temperature. And that a network of high centrality contacts connects distant regions of the protomer to the gate, serving as a means for the temperature-sensitive contact modes to allosterically regulate channel gating. Using a random forest model, we show that the contact states of specific temperature-sensitive modes are indeed predictive of the channel gate's state. Supporting the physical validity of these modes and networks are several residues identified with our analyses that are reported in literature to be functionally critical. Our results offer high resolution insight into thermo-TRP channel function and demonstrate the utility of temperature-sensitive contact analysis.


Assuntos
Repetição de Anquirina , Temperatura , Subunidades Proteicas/química
13.
Biochem Biophys Res Commun ; 683: 149077, 2023 11 26.
Artigo em Inglês | MEDLINE | ID: mdl-37890200

RESUMO

Targeted cytokine delivery has been gaining popularity in cancer immunotherapy since systemic recombinant cytokine treatment has not been successful due to low response rate and systemic toxicities in the clinical studies. In order to address these issues, we propose a new concept that cytokine signal is specifically activated at tumor-micro-environment (TME) by delivering two protein subunits of heterodimeric cytokine fused with a tumor targeting antibody respectively to TME and by bridging the two subunits into active heterodimeric form.Interleukin-12 (IL-12) is one of the major cytokines which can induce immune activation. IL-12 consists of two protein subunits which are p35 and p40. IL-12 signaling is initiated when it forms as the heterodimeric protein and binds to IL-12 receptor complex. We made fusion proteins of both IL-12p35 and IL-12p40 targeting specific tumor associated antigens (TAAs) and demonstrated the formation of bioactive IL12p70 with TME targeting antibody toward both p35 and p40 to form as the active molecule. We describe our concept validation in an in vitro based functional assay.


Assuntos
Citocinas , Neoplasias , Humanos , Subunidades Proteicas , Interleucina-12 , Proteínas Recombinantes , Neoplasias/terapia , Subunidade p40 da Interleucina-12 , Microambiente Tumoral
14.
Int J Mol Sci ; 24(20)2023 Oct 17.
Artigo em Inglês | MEDLINE | ID: mdl-37894940

RESUMO

Single-nucleotide polymorphisms in G protein subunits are linked to an increased risk of cardiovascular events among the general population. We assessed the effects of GNB3 c.825C > T, GNAQ -695/-694GC > TT, and GNAS c.393C > T polymorphisms on the risk of cardiovascular events among 454 patients undergoing renal replacement therapy. The patients were followed up for a median of 4.5 years after the initiation of dialysis. Carriers of the TT/TT genotype of GNAQ required stenting because of coronary artery stenosis (p = 0.0009) and developed cardiovascular events involving more than one organ system (p = 0.03) significantly earlier and more frequently than did the GC/TT or GC/GC genotypes. Multivariate analysis found that the TT/TT genotype of GNAQ was an independent risk factor for coronary artery stenosis requiring stent (hazard ratio, 4.5; p = 0.001), cardiovascular events (hazard ratio, 1.93; p = 0.04) and cardiovascular events affecting multiple organs (hazard ratio, 4.9; p = 0.03). In the subgroup of male patients left ventricular dilatation with abnormally increased LVEDD values occurred significantly more frequently in TT genotypes of GNB3 than in CT/CC genotypes (p = 0.007). Our findings suggest that male dialysis patients carrying the TT genotype of GNB3 are at higher risk of left ventricular dilatation and that dialysis patients carrying the TT/TT genotype of GNAQ are prone to coronary artery stenosis and severe cardiovascular events.


Assuntos
Estenose Coronária , Proteínas Heterotriméricas de Ligação ao GTP , Humanos , Masculino , Genótipo , Subunidades alfa Gq-G11 de Proteínas de Ligação ao GTP/genética , Proteínas Heterotriméricas de Ligação ao GTP/genética , Polimorfismo de Nucleotídeo Único , Subunidades Proteicas/genética , Diálise Renal/efeitos adversos , Terapia de Substituição Renal , Feminino
15.
Xi Bao Yu Fen Zi Mian Yi Xue Za Zhi ; 39(10): 946-951, 2023 Oct.
Artigo em Chinês | MEDLINE | ID: mdl-37882719

RESUMO

Since the outbreak of corona virus disease 2019 (COVID-19), viral strains have mutated and evolved. Vaccine research is the most direct and effective way to control COVID-19. According to different production mechanisms, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) vaccines included inactivated virus vaccine, live attenuated vaccine, mRNA vaccine, DNA vaccine, viral vector vaccine, virus-like particle vaccine and protein subunit vaccine. Among them, viral protein subunit vaccine has a wide application prospect due to its high safety and effectiveness. Viral nucleocapsid protein has high immunogenicity and low variability which could be a new direction for vaccine production. We summarized the current development of vaccine research by reviewing the current progress, vaccine safety and vaccine immune efficiency. It is hoped that the proposed possible development strategies could provide a reference for epidemic prevention work in future.


Assuntos
COVID-19 , Vacinas de DNA , Humanos , SARS-CoV-2/genética , COVID-19/prevenção & controle , Subunidades Proteicas , Proteínas do Nucleocapsídeo
16.
J Biol Chem ; 299(11): 105285, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37742920

RESUMO

Photoactivated adenylate cyclases (PACs) are multidomain BLUF proteins that regulate the cellular levels of cAMP in a light-dependent manner. The signaling route and dynamics of PAC from Oscillatoria acuminata (OaPAC), which consists of a light sensor BLUF domain, an adenylate cyclase domain, and a connector helix (α3-helix), were studied by detecting conformational changes in the protein moiety. Although circular dichroism and small-angle X-ray scattering measurements did not show significant changes upon light illumination, the transient grating method successfully detected light-induced changes in the diffusion coefficient (diffusion-sensitive conformational change (DSCC)) of full-length OaPAC and the BLUF domain with the α3-helix. DSCC of full-length OaPAC was observed only when both protomers in a dimer were photoconverted. This light intensity dependence suggests that OaPAC is a cyclase with a nonlinear light intensity response. The enzymatic activity indeed nonlinearly depends on light intensity, that is, OaPAC is activated under strong light conditions. It was also found that both DSCC and enzymatic activity were suppressed by a mutation in the W90 residue, indicating the importance of the highly conserved Trp in many BLUF domains for the function. Based on these findings, a reaction scheme was proposed together with the reaction dynamics.


Assuntos
Adenilil Ciclases , Proteínas de Bactérias , Luz , Transdução de Sinais , Adenilil Ciclases/genética , Adenilil Ciclases/metabolismo , Adenilil Ciclases/efeitos da radiação , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Proteínas de Bactérias/efeitos da radiação , Subunidades Proteicas , Ativação Enzimática/efeitos da radiação , Mutação
17.
Front Immunol ; 14: 1238586, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37654500

RESUMO

Tuberculosis (TB), also known as the "White Plague", is caused by Mycobacterium tuberculosis (Mtb). Before the COVID-19 epidemic, TB had the highest mortality rate of any single infectious disease. Vaccination is considered one of the most effective strategies for controlling TB. Despite the limitations of the Bacille Calmette-Guérin (BCG) vaccine in terms of protection against TB among adults, it is currently the only licensed TB vaccine. Recently, with the evolution of bioinformatics and structural biology techniques to screen and optimize protective antigens of Mtb, the tremendous potential of protein subunit vaccines is being exploited. Multistage subunit vaccines obtained by fusing immunodominant antigens from different stages of TB infection are being used both to prevent and to treat TB. Additionally, the development of novel adjuvants is compensating for weaknesses of immunogenicity, which is conducive to the flourishing of subunit vaccines. With advances in the development of animal models, preclinical vaccine protection assessments are becoming increasingly accurate. This review summarizes progress in the research of protein subunit TB vaccines during the past decades to facilitate the further optimization of protein subunit vaccines that may eradicate TB.


Assuntos
COVID-19 , Vacinas contra a Tuberculose , Tuberculose , Animais , Subunidades Proteicas , COVID-19/prevenção & controle , Vacinas de Subunidades , Tuberculose/prevenção & controle , Vacina BCG
18.
Nat Chem ; 15(12): 1664-1671, 2023 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-37667012

RESUMO

Molecular systems with coincident cyclic and superhelical symmetry axes have considerable advantages for materials design as they can be readily lengthened or shortened by changing the length of the constituent monomers. Among proteins, alpha-helical coiled coils have such symmetric, extendable architectures, but are limited by the relatively fixed geometry and flexibility of the helical protomers. Here we describe a systematic approach to generating modular and rigid repeat protein oligomers with coincident C2 to C8 and superhelical symmetry axes that can be readily extended by repeat propagation. From these building blocks, we demonstrate that a wide range of unbounded fibres can be systematically designed by introducing hydrophilic surface patches that force staggering of the monomers; the geometry of such fibres can be precisely tuned by varying the number of repeat units in the monomer and the placement of the hydrophilic patches.


Assuntos
Nanofibras , Modelos Moleculares , Conformação Proteica em alfa-Hélice , Subunidades Proteicas
19.
Nature ; 622(7981): 195-201, 2023 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-37730991

RESUMO

Type A γ-aminobutyric acid receptors (GABAARs) are the principal inhibitory receptors in the brain and the target of a wide range of clinical agents, including anaesthetics, sedatives, hypnotics and antidepressants1-3. However, our understanding of GABAAR pharmacology has been hindered by the vast number of pentameric assemblies that can be derived from 19 different subunits4 and the lack of structural knowledge of clinically relevant receptors. Here, we isolate native murine GABAAR assemblies containing the widely expressed α1 subunit and elucidate their structures in complex with drugs used to treat insomnia (zolpidem (ZOL) and flurazepam) and postpartum depression (the neurosteroid allopregnanolone (APG)). Using cryo-electron microscopy (cryo-EM) analysis and single-molecule photobleaching experiments, we uncover three major structural populations in the brain: the canonical α1ß2γ2 receptor containing two α1 subunits, and two assemblies containing one α1 and either an α2 or α3 subunit, in which the single α1-containing receptors feature a more compact arrangement between the transmembrane and extracellular domains. Interestingly, APG is bound at the transmembrane α/ß subunit interface, even when not added to the sample, revealing an important role for endogenous neurosteroids in modulating native GABAARs. Together with structurally engaged lipids, neurosteroids produce global conformational changes throughout the receptor that modify the ion channel pore and the binding sites for GABA and insomnia medications. Our data reveal the major α1-containing GABAAR assemblies, bound with endogenous neurosteroid, thus defining a structural landscape from which subtype-specific drugs can be developed.


Assuntos
Microscopia Crioeletrônica , Neuroesteroides , Receptores de GABA-A , Ácido gama-Aminobutírico , Animais , Camundongos , Sítios de Ligação/efeitos dos fármacos , Depressão Pós-Parto/tratamento farmacológico , Flurazepam/farmacologia , Ácido gama-Aminobutírico/metabolismo , Hipnóticos e Sedativos/farmacologia , Ativação do Canal Iônico/efeitos dos fármacos , Neuroesteroides/metabolismo , Neuroesteroides/farmacologia , Fotodegradação , Pregnanolona/farmacologia , Conformação Proteica/efeitos dos fármacos , Subunidades Proteicas/química , Subunidades Proteicas/efeitos dos fármacos , Subunidades Proteicas/metabolismo , Receptores de GABA-A/química , Receptores de GABA-A/efeitos dos fármacos , Receptores de GABA-A/metabolismo , Receptores de GABA-A/ultraestrutura , Distúrbios do Início e da Manutenção do Sono/tratamento farmacológico , Zolpidem/farmacologia
20.
Phys Chem Chem Phys ; 25(37): 25603-25618, 2023 Sep 27.
Artigo em Inglês | MEDLINE | ID: mdl-37721108

RESUMO

Near-edge X-ray absorption mass spectrometry (NEXAMS) around the nitrogen and oxygen K-edges was employed on gas-phase peptides to probe the electronic transitions related to their protonation sites, namely at basic side chains, the N-terminus and the amide oxygen. The experimental results are supported by replica exchange molecular dynamics and density-functional theory and restricted open-shell configuration with single calculations to attribute the transitions responsible for the experimentally observed resonances. We studied five tailor-made glycine-based pentapeptides, where we identified the signature of the protonation site of N-terminal proline, histidine, lysine and arginine, at 406 eV, corresponding to N 1s → σ*(NHx+) (x = 2 or 3) transitions, depending on the peptides. We compared the spectra of pentaglycine and triglycine to evaluate the sensitivity of NEXAMS to protomers. Separate resonances have been identified to distinguish two protomers in triglycine, the protonation site at the N-terminus at 406 eV and the protonation site at the amide oxygen characterized by a transition at 403.1 eV.


Assuntos
Amidas , Peptídeos , Eletrônica , Ácido Nitrilotriacético , Oxigênio , Subunidades Proteicas , Raios X
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