Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 5.518
Filtrar
1.
Sheng Wu Gong Cheng Xue Bao ; 36(8): 1536-1545, 2020 Aug 25.
Artigo em Chinês | MEDLINE | ID: mdl-32924352

RESUMO

Moschus chrysogaster (sifanicus) viral hemorrhagic disease (McVHD) is an acute and highly lethal infectious disease caused by Moschus chrysogaster hemorrhagic disease virus (McHDV) whose genome sequence is highly homologous with rabbit hemorrhagic disease virus. To screen the protective antigen of McHDV and set the basis for study of McVHD vaccine, the antigen epitope of major structural protein VP60 of McHDV was analyzed, and the specific primers were designed to obtain three amplified DNA sequences encoding the main antigen epitope of VP60 from McHDV by using RT-PCR. Then the three DNA fragments were sequenced and cloned to prokaryotic expression vector with pET-28a(+) by using overlap extension PCR, and finally the prokaryotic expression plasmid pET-truncated-VP60 was constructed. Subsequently, the pET-truncated-VP60 was transformed into Escherichia coli BL21(DE3), and the recombinant proteins were expressed by IPTG induction. Finally, the expressed protein was purified and applied to immunize that without immunizing with RHD vaccine, then the antiserum titers were evaluated by the hemagglutination inhibition test, and the immune-protective efficacy of the recombinant proteins was observed and analyzed through animal challenge test. The results showed that the multi-epitope DNA fragments of VP60 of McHDV was successfully expressed in the form of inclusion bodies in E. coli, and the relative molecular weight of recombinant proteins is about 45 kDa. After immunized with the recombinant proteins, 100% of New Zealand white rabbits were resistant to attack of McHDV, which indicates efficient immune-protective efficacy of chosen epitope recombinant protein. The study laid a foundation for the development of the new subunit vaccines of McVHD.


Assuntos
Infecções por Caliciviridae , Expressão Gênica , Vírus da Doença Hemorrágica de Coelhos , Proteínas Estruturais Virais , Animais , Infecções por Caliciviridae/imunologia , Infecções por Caliciviridae/virologia , Epitopos/genética , Escherichia coli/genética , Coelhos , Proteínas Estruturais Virais/genética , Proteínas Estruturais Virais/metabolismo
2.
PLoS Pathog ; 16(8): e1008753, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32866207

RESUMO

The induction of broad and potent immunity by vaccines is the key focus of research efforts aimed at protecting against HIV-1 infection. Soluble native-like HIV-1 envelope glycoproteins have shown promise as vaccine candidates as they can induce potent autologous neutralizing responses in rabbits and non-human primates. In this study, monoclonal antibodies were isolated and characterized from rhesus macaques immunized with the BG505 SOSIP.664 trimer to better understand vaccine-induced antibody responses. Our studies reveal a diverse landscape of antibodies recognizing immunodominant strain-specific epitopes and non-neutralizing neo-epitopes. Additionally, we isolated a subset of mAbs against an epitope cluster at the gp120-gp41 interface that recognize the highly conserved fusion peptide and the glycan at position 88 and have characteristics akin to several human-derived broadly neutralizing antibodies.


Assuntos
Vacinas contra a AIDS/imunologia , Mapeamento de Epitopos , Epitopos/imunologia , Anticorpos Anti-HIV/imunologia , Proteína gp120 do Envelope de HIV/imunologia , Proteína gp41 do Envelope de HIV/imunologia , HIV-1/imunologia , Vacinas contra a AIDS/genética , Animais , Anticorpos Monoclonais Murinos/imunologia , Epitopos/genética , Anticorpos Anti-HIV/genética , Proteína gp41 do Envelope de HIV/genética , HIV-1/genética , Macaca mulatta , Multimerização Proteica/genética , Multimerização Proteica/imunologia
3.
Cell Mol Immunol ; 17(10): 1095-1097, 2020 10.
Artigo em Inglês | MEDLINE | ID: mdl-32895485
4.
Genome Med ; 12(1): 70, 2020 08 13.
Artigo em Inglês | MEDLINE | ID: mdl-32791978

RESUMO

BACKGROUND: The ongoing COVID-19 pandemic has created an urgency to identify novel vaccine targets for protective immunity against SARS-CoV-2. Early reports identify protective roles for both humoral and cell-mediated immunity for SARS-CoV-2. METHODS: We leveraged our bioinformatics binding prediction tools for human leukocyte antigen (HLA)-I and HLA-II alleles that were developed using mass spectrometry-based profiling of individual HLA-I and HLA-II alleles to predict peptide binding to diverse allele sets. We applied these binding predictors to viral genomes from the Coronaviridae family and specifically focused on T cell epitopes from SARS-CoV-2 proteins. We assayed a subset of these epitopes in a T cell induction assay for their ability to elicit CD8+ T cell responses. RESULTS: We first validated HLA-I and HLA-II predictions on Coronaviridae family epitopes deposited in the Virus Pathogen Database and Analysis Resource (ViPR) database. We then utilized our HLA-I and HLA-II predictors to identify 11,897 HLA-I and 8046 HLA-II candidate peptides which were highly ranked for binding across 13 open reading frames (ORFs) of SARS-CoV-2. These peptides are predicted to provide over 99% allele coverage for the US, European, and Asian populations. From our SARS-CoV-2-predicted peptide-HLA-I allele pairs, 374 pairs identically matched what was previously reported in the ViPR database, originating from other coronaviruses with identical sequences. Of these pairs, 333 (89%) had a positive HLA binding assay result, reinforcing the validity of our predictions. We then demonstrated that a subset of these highly predicted epitopes were immunogenic based on their recognition by specific CD8+ T cells in healthy human donor peripheral blood mononuclear cells (PBMCs). Finally, we characterized the expression of SARS-CoV-2 proteins in virally infected cells to prioritize those which could be potential targets for T cell immunity. CONCLUSIONS: Using our bioinformatics platform, we identify multiple putative epitopes that are potential targets for CD4+ and CD8+ T cells, whose HLA binding properties cover nearly the entire population. We also confirm that our binding predictors can predict epitopes eliciting CD8+ T cell responses from multiple SARS-CoV-2 proteins. Protein expression and population HLA allele coverage, combined with the ability to identify T cell epitopes, should be considered in SARS-CoV-2 vaccine design strategies and immune monitoring.


Assuntos
Infecções por Coronavirus/imunologia , Epitopos/imunologia , Antígenos HLA/imunologia , Pneumonia Viral/imunologia , Linfócitos T/imunologia , Vacinas Virais/imunologia , Alelos , Afinidade de Anticorpos , Biologia Computacional , Infecções por Coronavirus/genética , Infecções por Coronavirus/prevenção & controle , Epitopos/química , Epitopos/genética , Genoma Viral , Antígenos HLA/química , Antígenos HLA/genética , Humanos , Imunogenicidade da Vacina , Espectrometria de Massas , Pandemias , Vacinas Virais/química , Vacinas Virais/genética
5.
PLoS Pathog ; 16(8): e1008736, 2020 08.
Artigo em Inglês | MEDLINE | ID: mdl-32745149

RESUMO

Human cytomegalovirus (HCMV) is one of the main causative agents of congenital viral infection in neonates. HCMV infection also causes serious morbidity and mortality among organ transplant patients. Glycoprotein B (gB) is a major target for HCMV neutralizing antibodies, yet the underlying neutralization mechanisms remain largely unknown. Here we report that 3-25, a gB-specific monoclonal antibody previously isolated from a healthy HCMV-positive donor, efficiently neutralized 14 HCMV strains in both ARPE-19 cells and MRC-5 cells. The core epitope of 3-25 was mapped to a highly conserved linear epitope on antigenic domain 2 (AD-2) of gB. A 1.8 Å crystal structure of 3-25 Fab in complex with the peptide epitope revealed the molecular determinants of 3-25 binding to gB at atomic resolution. Negative-staining electron microscopy (EM) 3D reconstruction of 3-25 Fab in complex with de-glycosylated postfusion gB showed that 3-25 Fab fully occupied the gB trimer at the N-terminus with flexible binding angles. Functionally, 3-25 efficiently inhibited HCMV infection at a post-attachment step by interfering with viral membrane fusion, and restricted post-infection viral spreading in ARPE-19 cells. Interestingly, bivalency was required for HCMV neutralization by AD-2 specific antibody 3-25 but not the AD-4 specific antibody LJP538. In contrast, bivalency was not required for HCMV binding by both antibodies. Taken together, our results reveal the structural basis of gB recognition by 3-25 and demonstrate that inhibition of viral membrane fusion and a requirement of bivalency may be common for gB AD-2 specific neutralizing antibody.


Assuntos
Anticorpos Antivirais/imunologia , Infecções por Citomegalovirus/imunologia , Citomegalovirus/imunologia , Epitopos/imunologia , Proteínas do Envelope Viral/imunologia , Motivos de Aminoácidos , Anticorpos Neutralizantes/imunologia , Sequência Conservada , Citomegalovirus/química , Citomegalovirus/genética , Citomegalovirus/fisiologia , Infecções por Citomegalovirus/virologia , Epitopos/química , Epitopos/genética , Humanos , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/genética , Internalização do Vírus
6.
Med Hypotheses ; 143: 110151, 2020 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-32763661

RESUMO

Today it remains unclear why children seem to be less likely to get infected by COVID-19 or why they appear to be less symptomatic after infections. All individuals, especially children, are exposed to various viruses including human coronavirus (CoVs) that can generally lead to respiratory infections. We hypothesize that recurrent CoVs exposure may induce an effective antiviral B and T-cell-mediated adaptive immune response, which could also be protective against COVID-19. Based on the high-homology between the Spike protein epitopes of taxonomically-related coronaviruses, we theorize that past/recurrent contact with CoVs might shield children also against the circulating COVID-19 through a possible neutralizing antibody response previously CoVs-induced. This would open up possible lines of research for the development of live-attenuated virus vaccines from CoVs. Future research is desirable to confirm or disprove such hypothesis.


Assuntos
Betacoronavirus , Infecções por Coronavirus/epidemiologia , Memória Imunológica , Modelos Imunológicos , Pandemias , Pneumonia Viral/epidemiologia , Adulto , Distribuição por Idade , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/imunologia , Antígenos Virais/genética , Antígenos Virais/imunologia , Betacoronavirus/genética , Betacoronavirus/imunologia , Linfócitos T CD4-Positivos/imunologia , Criança , Coronavirus/genética , Coronavirus/imunologia , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Infecções por Coronavirus/virologia , Reações Cruzadas , Resistência à Doença , Epitopos/genética , Epitopos/imunologia , Humanos , Peptidil Dipeptidase A/análise , Pneumonia Viral/imunologia , Alvéolos Pulmonares/química , Receptores Virais/análise , Infecções Respiratórias/epidemiologia , Infecções Respiratórias/imunologia , Infecções Respiratórias/virologia , Homologia de Sequência de Aminoácidos , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia , Vacinas Atenuadas , Vacinas Virais
7.
Cell Syst ; 11(2): 131-144.e6, 2020 08 26.
Artigo em Inglês | MEDLINE | ID: mdl-32721383

RESUMO

We present a combinatorial machine learning method to evaluate and optimize peptide vaccine formulations for SARS-CoV-2. Our approach optimizes the presentation likelihood of a diverse set of vaccine peptides conditioned on a target human-population HLA haplotype distribution and expected epitope drift. Our proposed SARS-CoV-2 MHC class I vaccine formulations provide 93.21% predicted population coverage with at least five vaccine peptide-HLA average hits per person (≥ 1 peptide: 99.91%) with all vaccine peptides perfectly conserved across 4,690 geographically sampled SARS-CoV-2 genomes. Our proposed MHC class II vaccine formulations provide 97.21% predicted coverage with at least five vaccine peptide-HLA average hits per person with all peptides having an observed mutation probability of ≤ 0.001. We provide an open-source implementation of our design methods (OptiVax), vaccine evaluation tool (EvalVax), as well as the data used in our design efforts here: https://github.com/gifford-lab/optivax.


Assuntos
Betacoronavirus/imunologia , Haplótipos , Antígenos de Histocompatibilidade Classe II/genética , Antígenos de Histocompatibilidade Classe I/genética , Análise de Sequência de DNA/métodos , Vacinas de Subunidades/imunologia , Vacinas Virais/imunologia , Betacoronavirus/genética , Infecções por Coronavirus/genética , Infecções por Coronavirus/imunologia , Infecções por Coronavirus/prevenção & controle , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Antígenos de Histocompatibilidade Classe I/química , Antígenos de Histocompatibilidade Classe I/imunologia , Antígenos de Histocompatibilidade Classe II/química , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Aprendizado de Máquina , Análise de Sequência de DNA/normas , Vacinas de Subunidades/química , Vacinas de Subunidades/genética , Vacinas Virais/química , Vacinas Virais/genética
8.
BMC Infect Dis ; 20(1): 493, 2020 Jul 10.
Artigo em Inglês | MEDLINE | ID: mdl-32650739

RESUMO

BACKGROUND: Toxoplasma gondii is an obligate intracellular parasite that can infect almost all warm-blooded animals, avian species and humans. Toxoplasmosis is asymptomatic in healthy individuals, whereas it may lead to death in immune suppressed or deficient patients. A vaccine against T. gondii is required to prevent consequences of the infection. The aim of this study is to generate a multivalent recombinant protein vaccine against T. gondii. METHODS: 49 previously discovered antigenic proteins of T gondii were evaluated by their expression level in E. coli and by comprehensive bioinformatics analyses to determine antigenic epitopes. Based on these analyses, six vaccine candidate proteins were selected to generate a hexavalent recombinant protein vaccine adjuvanted with Montanide ISA 50 V. Humoral and cellular immune responses were determined by flow cytometry and ELISA. Vaccinated mice were challenged with T. gondii Ankara strain tachyzoites. RESULTS: In mice vaccinated with hexavalent vaccine, strong total IgG (P < 0.0001) and IgG2a (P < 0.001) responses were induced compared to controls, the ratio of CD4+ and CD8+ T lymphocytes secreting IFN-γ increased, and significantly higher extracellular IFN-γ secretion was achieved compared to the controls (P < 0.001). The survival time of the vaccinated mice increased to 8.38 ± 2.13 days which was significantly higher than controls (P < 0.01). CONCLUSIONS: Altogether, these results show that the hexavalent vaccine which is developed for the first time against T. gondii induced strong and balanced Th1 and Th2 immune responses as well as conferred significant protection against challenge with lethal toxoplasmosis in murine model.


Assuntos
Adjuvantes Imunológicos/farmacologia , Vacinas Protozoárias/farmacologia , Toxoplasmose/prevenção & controle , Vacinas de DNA/farmacologia , Animais , Ensaio de Imunoadsorção Enzimática , Epitopos/genética , Epitopos/imunologia , Escherichia coli/genética , Feminino , Imunidade Celular/efeitos dos fármacos , Imunidade Humoral/efeitos dos fármacos , Imunoglobulina G/sangue , Camundongos , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Vacinas Protozoárias/genética , Vacinas Protozoárias/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Toxoplasma/patogenicidade , Toxoplasmose/imunologia , Vacinas de DNA/imunologia
9.
Viruses ; 12(6)2020 06 03.
Artigo em Inglês | MEDLINE | ID: covidwho-532726

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe respiratory illness in humans; the second-largest and most deadly outbreak to date occurred in Saudi Arabia. The dromedary camel is considered a possible host of the virus and also to act as a reservoir, transmitting the virus to humans. Here, we studied evolutionary relationships for 31 complete genomes of betacoronaviruses, including eight newly sequenced MERS-CoV genomes isolated from dromedary camels in Saudi Arabia. Through bioinformatics tools, we also used available sequences and 3D structure of MERS-CoV spike glycoprotein to predict MERS-CoV epitopes and assess antibody binding affinity. Phylogenetic analysis showed the eight new sequences have close relationships with existing strains detected in camels and humans in Arabian Gulf countries. The 2019-nCov strain appears to have higher homology to both bat coronavirus and SARS-CoV than to MERS-CoV strains. The spike protein tree exhibited clustering of MERS-CoV sequences similar to the complete genome tree, except for one sequence from Qatar (KF961222). B cell epitope analysis determined that the MERS-CoV spike protein has 24 total discontinuous regions from which just six epitopes were selected with score values of >80%. Our results suggest that the virus circulates by way of camels crossing the borders of Arabian Gulf countries. This study contributes to finding more effective vaccines in order to provide long-term protection against MERS-CoV and identifying neutralizing antibodies.


Assuntos
Camelus/virologia , Infecções por Coronavirus/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Glicoproteína da Espícula de Coronavírus/genética , Sequência de Aminoácidos , Animais , Betacoronavirus/classificação , Betacoronavirus/genética , Betacoronavirus/isolamento & purificação , Evolução Biológica , DNA Complementar/química , DNA Viral/química , Epitopos/análise , Epitopos/química , Epitopos/genética , Biblioteca Gênica , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/classificação , Coronavírus da Síndrome Respiratória do Oriente Médio/isolamento & purificação , Filogenia , RNA Viral/análise , RNA Viral/química , RNA Viral/isolamento & purificação , Arábia Saudita
10.
Nat Commun ; 11(1): 2841, 2020 06 05.
Artigo em Inglês | MEDLINE | ID: mdl-32503989

RESUMO

The capsid of human papillomavirus (HPV) spontaneously arranges into a T = 7 icosahedral particle with 72 L1 pentameric capsomeres associating via disulfide bonds between Cys175 and Cys428. Here, we design a capsomere-hybrid virus-like particle (chVLP) to accommodate multiple types of L1 pentamers by the reciprocal assembly of single C175A and C428A L1 mutants, either of which alone encumbers L1 pentamer particle self-assembly. We show that co-assembly between any pair of C175A and C428A mutants across at least nine HPV genotypes occurs at a preferred equal molar stoichiometry, irrespective of the type or number of L1 sequences. A nine-valent chVLP vaccine-formed through the structural clustering of HPV epitopes-confers neutralization titers that are comparable with that of Gardasil 9 and elicits minor cross-neutralizing antibodies against some heterologous HPV types. These findings may pave the way for a new vaccine design that targets multiple pathogenic variants or cancer cells bearing diverse neoantigens.


Assuntos
Proteínas do Capsídeo/imunologia , Neoplasias/terapia , Papillomaviridae/imunologia , Infecções por Papillomavirus/terapia , Vacinas contra Papillomavirus/imunologia , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Proteínas do Capsídeo/administração & dosagem , Proteínas do Capsídeo/genética , Desenho de Fármacos , Epitopos/genética , Epitopos/imunologia , Feminino , Humanos , Imunogenicidade da Vacina , Camundongos , Modelos Animais , Mutação , Neoplasias/virologia , Testes de Neutralização , Papillomaviridae/genética , Infecções por Papillomavirus/virologia , Vacinas contra Papillomavirus/administração & dosagem , Vacinas contra Papillomavirus/genética , Multimerização Proteica/genética , Multimerização Proteica/imunologia , Vacinas de Partículas Semelhantes a Vírus/administração & dosagem , Vacinas de Partículas Semelhantes a Vírus/genética , Vacinas de Partículas Semelhantes a Vírus/imunologia
11.
Proc Natl Acad Sci U S A ; 117(26): 15148-15159, 2020 06 30.
Artigo em Inglês | MEDLINE | ID: mdl-32541028

RESUMO

The potency of adoptive T cell therapies targeting the cell surface antigen CD19 has been demonstrated in hematopoietic cancers. It has been difficult to identify appropriate targets in nonhematopoietic tumors, but one class of antigens that have shown promise is aberrant O-glycoprotein epitopes. It has long been known that dysregulated synthesis of O-linked (threonine or serine) sugars occurs in many cancers, and that this can lead to the expression of cell surface proteins containing O-glycans comprised of a single N-acetylgalactosamine (GalNAc, known as Tn antigen) rather than the normally extended carbohydrate. Previously, we used the scFv fragment of antibody 237 as a chimeric antigen receptor (CAR) to mediate recognition of mouse tumor cells that bear its cognate Tn-glycopeptide epitope in podoplanin, also called OTS8. Guided by the structure of the 237 Fab:Tn-OTS8-glycopeptide complex, here we conducted a deep mutational scan showing that residues flanking the Tn-glycan contributed significant binding energy to the interaction. Design of 237-scFv libraries in the yeast display system allowed us to isolate scFv variants with higher affinity for Tn-OTS8. Selection with a noncognate human antigen, Tn-MUC1, yielded scFv variants that were broadly reactive with multiple Tn-glycoproteins. When configured as CARs, engineered T cells expressing these scFv variants showed improved activity against mouse and human cancer cell lines defective in O-linked glycosylation. This strategy provides CARs with Tn-peptide specificities, all based on a single scFv scaffold, that allows the same CAR to be tested for toxicity in mice and efficacy against mouse and human tumors.


Assuntos
Antígenos Glicosídicos Associados a Tumores/imunologia , Receptores de Antígenos Quiméricos/metabolismo , Linfócitos T/fisiologia , Sequência de Aminoácidos , Animais , Anticorpos , Linhagem Celular Tumoral , Evolução Molecular Direcionada , Epitopos/genética , Humanos , Camundongos , Modelos Moleculares , Mutação , Conformação Proteica , Receptores de Antígenos Quiméricos/genética
12.
Viruses ; 12(6)2020 06 03.
Artigo em Inglês | MEDLINE | ID: mdl-32503352

RESUMO

Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe respiratory illness in humans; the second-largest and most deadly outbreak to date occurred in Saudi Arabia. The dromedary camel is considered a possible host of the virus and also to act as a reservoir, transmitting the virus to humans. Here, we studied evolutionary relationships for 31 complete genomes of betacoronaviruses, including eight newly sequenced MERS-CoV genomes isolated from dromedary camels in Saudi Arabia. Through bioinformatics tools, we also used available sequences and 3D structure of MERS-CoV spike glycoprotein to predict MERS-CoV epitopes and assess antibody binding affinity. Phylogenetic analysis showed the eight new sequences have close relationships with existing strains detected in camels and humans in Arabian Gulf countries. The 2019-nCov strain appears to have higher homology to both bat coronavirus and SARS-CoV than to MERS-CoV strains. The spike protein tree exhibited clustering of MERS-CoV sequences similar to the complete genome tree, except for one sequence from Qatar (KF961222). B cell epitope analysis determined that the MERS-CoV spike protein has 24 total discontinuous regions from which just six epitopes were selected with score values of >80%. Our results suggest that the virus circulates by way of camels crossing the borders of Arabian Gulf countries. This study contributes to finding more effective vaccines in order to provide long-term protection against MERS-CoV and identifying neutralizing antibodies.


Assuntos
Camelus/virologia , Infecções por Coronavirus/virologia , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Glicoproteína da Espícula de Coronavírus/genética , Sequência de Aminoácidos , Animais , Betacoronavirus/classificação , Betacoronavirus/genética , Betacoronavirus/isolamento & purificação , Evolução Biológica , DNA Complementar/química , DNA Viral/química , Epitopos/análise , Epitopos/química , Epitopos/genética , Biblioteca Gênica , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/classificação , Coronavírus da Síndrome Respiratória do Oriente Médio/isolamento & purificação , Filogenia , RNA Viral/análise , RNA Viral/química , RNA Viral/isolamento & purificação , Arábia Saudita
13.
Adv Exp Med Biol ; 1194: 359-371, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32468552

RESUMO

Monoclonal antibodies (mAbs) constitute a promising class of therapeutics, since ca. 25% of all biotech drugs in development are mAbs. Even though their therapeutic value is now well established, human- and murine-derived mAbs do have deficiencies, such as short in vivo lifespan and low stability. However, the most difficult obstacle to overcome, toward the exploitation of mAbs for disease treatment, is the prevention of the formation of protein aggregates. ANTISOMA is a pipeline for the reduction of the aggregation tendency of mAbs through the decrease in their intrinsic aggregation propensity, based on an automated amino acid substitution approach. The method takes into consideration the special features of mAbs and aims at proposing specific point mutations that could lead to the redesign of those promising therapeutics, without affecting their epitope-binding ability. The method is available online at http://bioinformatics.biol.uoa.gr/ANTISOMA .


Assuntos
Anticorpos Monoclonais , Biologia Computacional , Agregação Patológica de Proteínas , Animais , Anticorpos Monoclonais/genética , Anticorpos Monoclonais/metabolismo , Anticorpos Monoclonais/uso terapêutico , Biologia Computacional/métodos , Epitopos/genética , Humanos , Camundongos , Agregação Patológica de Proteínas/tratamento farmacológico
14.
Clin Immunol ; 215: 108426, 2020 06.
Artigo em Inglês | MEDLINE | ID: covidwho-115814

Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Pneumocystis carinii/imunologia , Pneumonia por Pneumocystis/imunologia , Pneumonia Viral/imunologia , Proteínas Associadas a Surfactantes Pulmonares/química , Surfactantes Pulmonares/química , Glicoproteína da Espícula de Coronavírus/química , Sequência de Aminoácidos , Anticorpos Antivirais/química , Anticorpos Antivirais/genética , Betacoronavirus/patogenicidade , Coronavirus Humano 229E/imunologia , Infecções por Coronavirus/genética , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/imunologia , Reações Cruzadas , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Oligopeptídeos/química , Oligopeptídeos/genética , Oligopeptídeos/imunologia , Pandemias , Pneumocystis carinii/patogenicidade , Pneumonia por Pneumocystis/genética , Pneumonia por Pneumocystis/patologia , Pneumonia por Pneumocystis/virologia , Pneumonia Viral/genética , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Ligação Proteica , Proteínas Associadas a Surfactantes Pulmonares/genética , Proteínas Associadas a Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/metabolismo , Homologia de Sequência de Aminoácidos , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
15.
Clin Immunol ; 215: 108426, 2020 06.
Artigo em Inglês | MEDLINE | ID: covidwho-72350

Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Pneumocystis carinii/imunologia , Pneumonia por Pneumocystis/imunologia , Pneumonia Viral/imunologia , Proteínas Associadas a Surfactantes Pulmonares/química , Surfactantes Pulmonares/química , Glicoproteína da Espícula de Coronavírus/química , Sequência de Aminoácidos , Anticorpos Antivirais/química , Anticorpos Antivirais/genética , Betacoronavirus/patogenicidade , Coronavirus Humano 229E/imunologia , Infecções por Coronavirus/genética , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/imunologia , Reações Cruzadas , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Oligopeptídeos/química , Oligopeptídeos/genética , Oligopeptídeos/imunologia , Pandemias , Pneumocystis carinii/patogenicidade , Pneumonia por Pneumocystis/genética , Pneumonia por Pneumocystis/patologia , Pneumonia por Pneumocystis/virologia , Pneumonia Viral/genética , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Ligação Proteica , Proteínas Associadas a Surfactantes Pulmonares/genética , Proteínas Associadas a Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/metabolismo , Homologia de Sequência de Aminoácidos , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
16.
Clin Immunol ; 215: 108426, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32311462

Assuntos
Betacoronavirus/imunologia , Infecções por Coronavirus/imunologia , Pneumocystis carinii/imunologia , Pneumonia por Pneumocystis/imunologia , Pneumonia Viral/imunologia , Proteínas Associadas a Surfactantes Pulmonares/química , Surfactantes Pulmonares/química , Glicoproteína da Espícula de Coronavírus/química , Sequência de Aminoácidos , Anticorpos Antivirais/química , Anticorpos Antivirais/genética , Betacoronavirus/patogenicidade , Coronavirus Humano 229E/imunologia , Infecções por Coronavirus/genética , Infecções por Coronavirus/patologia , Infecções por Coronavirus/virologia , Coronavirus Humano OC43/imunologia , Reações Cruzadas , Epitopos/química , Epitopos/genética , Epitopos/imunologia , Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Oligopeptídeos/química , Oligopeptídeos/genética , Oligopeptídeos/imunologia , Pandemias , Pneumocystis carinii/patogenicidade , Pneumonia por Pneumocystis/genética , Pneumonia por Pneumocystis/patologia , Pneumonia por Pneumocystis/virologia , Pneumonia Viral/genética , Pneumonia Viral/patologia , Pneumonia Viral/virologia , Ligação Proteica , Proteínas Associadas a Surfactantes Pulmonares/genética , Proteínas Associadas a Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/imunologia , Surfactantes Pulmonares/metabolismo , Homologia de Sequência de Aminoácidos , Glicoproteína da Espícula de Coronavírus/genética , Glicoproteína da Espícula de Coronavírus/imunologia
17.
Food Chem ; 322: 126711, 2020 Aug 30.
Artigo em Inglês | MEDLINE | ID: mdl-32283362

RESUMO

Jug r 1, the major allergen of walnut, triggers severe allergic reactions through epitopes. Hence, research on the efficient strategy for analyzing the linear epitopes of Jug r 1 are necessary. In this work, bioinformatics analysis was used to predict the linear epitopes of Jug r 1. Overlapping peptide synthesis was used to map linear epitopes. In vitro simulated gastrointestinal digestion and HPLC-MS/MS were used to identify digestion-resistant peptides. The results showed that six predicted linear epitopes were AA28-35, AA42-49, AA55-62, AA65-73, AA97-104, and AA109-121. AA16-30 and AA125-139 were identified by the sera of walnut allergic patients. Five digestion-resistant peptides were AA19-33, AA40-45, AA54-74, AA96-106, and AA117-137. The predicted results only included one of the linear epitopes identified by sera, while the digestion-resistant peptides covered all. Therefore, the digestion-resistant property of food allergens may be a promising direction for studying the linear epitopes of Jug r 1.


Assuntos
Alérgenos/química , Epitopos/química , Juglans/química , Peptídeos/química , Alérgenos/genética , Alérgenos/imunologia , Sequência de Aminoácidos , Mapeamento de Epitopos , Epitopos/genética , Epitopos/imunologia , Hipersensibilidade Alimentar/imunologia , Humanos , Juglans/genética , Juglans/imunologia , Nozes/química , Nozes/genética , Nozes/imunologia , Peptídeos/imunologia , Análise de Sequência , Espectrometria de Massas em Tandem
18.
PLoS Biol ; 18(4): e3000665, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32275651

RESUMO

The correct subcellular distribution of proteins establishes the complex morphology and function of neurons. Fluorescence microscopy techniques are invaluable to investigate subcellular protein distribution, but they suffer from the limited ability to efficiently and reliably label endogenous proteins with fluorescent probes. We developed ORANGE: Open Resource for the Application of Neuronal Genome Editing, which mediates targeted genomic integration of epitope tags in rodent dissociated neuronal culture, in organotypic slices, and in vivo. ORANGE includes a knock-in library for in-depth investigation of endogenous protein distribution, viral vectors, and a detailed two-step cloning protocol to develop knock-ins for novel targets. Using ORANGE with (live-cell) superresolution microscopy, we revealed the dynamic nanoscale organization of endogenous neurotransmitter receptors and synaptic scaffolding proteins, as well as previously uncharacterized proteins. Finally, we developed a mechanism to create multiple knock-ins in neurons, mediating multiplex imaging of endogenous proteins. Thus, ORANGE enables quantification of expression, distribution, and dynamics for virtually any protein in neurons at nanoscale resolution.


Assuntos
Sistemas CRISPR-Cas , Epitopos/genética , Edição de Genes/métodos , Neurônios/imunologia , Proteínas/genética , Animais , Células Cultivadas , Dependovirus/genética , Feminino , Técnicas de Introdução de Genes , Genes Reporter , Vetores Genéticos , Genoma , Proteínas de Fluorescência Verde/genética , Proteínas de Fluorescência Verde/metabolismo , Masculino , Camundongos Transgênicos , Microscopia de Fluorescência , Imagem Molecular/métodos , Neurônios/fisiologia , Técnicas de Cultura de Órgãos , Proteínas/imunologia , Proteínas/metabolismo , Ratos Wistar , Receptores de N-Metil-D-Aspartato/genética , Receptores de N-Metil-D-Aspartato/metabolismo , Análise Espaço-Temporal
19.
Proc Natl Acad Sci U S A ; 117(18): 9942-9951, 2020 05 05.
Artigo em Inglês | MEDLINE | ID: mdl-32321835

RESUMO

Genetic variants within complement factor H (CFH), a major alternative complement pathway regulator, are associated with the development of age-related macular degeneration (AMD) and other complementopathies. This is explained with the reduced binding of CFH or its splice variant factor H-like protein 1 (FHL-1) to self-ligands or altered self-ligands (e.g., malondialdehyde [MDA]-modified molecules) involved in homeostasis, thereby causing impaired complement regulation. Considering the critical role of CFH in inhibiting alternative pathway activation on MDA-modified surfaces, we performed an unbiased genome-wide search for genetic variants that modify the ability of plasma CFH to bind MDA in 1,830 individuals and characterized the mechanistic basis and the functional consequences of this. In a cohort of healthy individuals, we identified rs1061170 in CFH and the deletion of CFHR3 and CFHR1 as dominant genetic variants that modify CFH/FHL-1 binding to MDA. We further demonstrated that FHR1 and FHR3 compete with CFH for binding to MDA-epitopes and that FHR1 displays the highest affinity toward MDA-epitopes compared to CFH and FHR3. Moreover, FHR1 bound to MDA-rich areas on necrotic cells and prevented CFH from mediating its cofactor activity on MDA-modified surfaces, resulting in enhanced complement activation. These findings provide a mechanistic explanation as to why the deletion of CFHR3 and CFHR1 is protective in AMD and highlight the importance of genetic variants within the CFH/CFHR3/CFHR1 locus in the recognition of altered-self in tissue homeostasis.


Assuntos
Proteínas Sanguíneas/genética , Proteínas Inativadoras do Complemento C3b/genética , Degeneração Macular/genética , Idoso , Fator H do Complemento/genética , Epitopos/genética , Feminino , Predisposição Genética para Doença , Estudo de Associação Genômica Ampla , Humanos , Degeneração Macular/patologia , Masculino , Malondialdeído/metabolismo , Pessoa de Meia-Idade , Polimorfismo de Nucleotídeo Único/genética , Ligação Proteica
20.
Methods Mol Biol ; 2131: 39-145, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32162250

RESUMO

The aim of this study was to use IEDB software to predict the suitable MERS-CoV epitope vaccine against the most known world population alleles through four selecting proteins such as S glycoprotein and envelope protein and their modification sequences after the pandemic spread of MERS-CoV in 2012. IEDB services is one of the computational methods; the output of this study showed that S glycoprotein, envelope (E) protein, and S and E protein modified sequences of MERS-CoV might be considered as a protective immunogenic with high conservancy because they can elect both neutralizing antibodies and T-cell responses when reacting with B-cell, T-helper cell, and cytotoxic T lymphocyte. NetCTL, NetChop, and MHC-NP were used to confirm our results. Population coverage analysis showed that the putative helper T-cell epitopes and CTL epitopes could cover most of the world population in more than 60 geographical regions. According to AllerHunter results, all those selected different protein showed non-allergen; this finding makes this computational vaccine study more desirable for vaccine synthesis.


Assuntos
Anticorpos Antivirais/imunologia , Infecções por Coronavirus/prevenção & controle , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Software , Glicoproteína da Espícula de Coronavírus/imunologia , Proteínas do Envelope Viral/imunologia , Vacinas Virais/imunologia , Anticorpos Neutralizantes/imunologia , Linfócitos B/imunologia , Infecções por Coronavirus/virologia , Bases de Dados Genéticas , Epitopos/genética , Epitopos/imunologia , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Mutação , Peptídeos/genética , Peptídeos/imunologia , Glicoproteína da Espícula de Coronavírus/genética , Linfócitos T Citotóxicos/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Vacinas Sintéticas/imunologia , Proteínas do Envelope Viral/genética
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA