Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 2.647
Filtrar
1.
J Chromatogr A ; 1643: 462076, 2021 Apr 26.
Artigo em Inglês | MEDLINE | ID: mdl-33789193

RESUMO

The present research is focused on the preliminary evaluation, in particular in relation to the advisable operational conditions, of a novel low duty cycle flow modulator. In such a respect, a fast comprehensive two-dimensional gas chromatography-mass spectrometry method is herein proposed. Applications on a C7-C30 series of alkanes, 64 fragrance allergens (plus 2 internal standards), and 5 perfumes, were carried out by using two different column sets, low-polarity + medium-polarity and low-polarity + low-polarity. In both cases, the first column was of dimensions 10 m × 0.25 mm ID × 0.25 µm df, while the second one was of dimensions 1 m × 0.10 mm ID × 0.10 µm df. A modulation period of 700 ms, with a re-injection period of 80 ms, was used in order to obtain a higher duty cycle (measured to be approx. 0.04). Absolute quantification of the allergens was carried out by using two internal standards, namely 1,4-dibromobenzene and 4,4'-dibromobiphenyl. In terms of limits of quantification the instrumental response was characterized by a wide variability, ranging between 9 ppb and 5.4 ppm for both column sets. A total number of 97 fragrance allergens were identified and quantified in five commercial perfumes.


Assuntos
Alérgenos/análise , Cromatografia Gasosa/métodos , Perfumes/química , Alcanos/análise , Alcanos/química , Alérgenos/química , Limite de Detecção , Perfumes/normas , Padrões de Referência , Espectrometria de Massas por Ionização por Electrospray
2.
Food Chem ; 346: 128953, 2021 Jun 01.
Artigo em Inglês | MEDLINE | ID: mdl-33412487

RESUMO

Tartary buckwheat is widely accepted as its nutritionalvalue. Some allergic reactions hinder its utilization. This research focused on evaluating the core epitope of 16 kDa allergen (Fag t 2) in tartary buckwheat. Six B- and seven T cell epitopes of Fag t 2 were predicted, and six B cell epitope-mutants were expressed in Pichia pastoris. Bioinformatics analysis and SDS-PAGE demonstrated that the molecular weight, isoelectric point and spatial structures of six mutant allergens were similar with Fag t 2, with the same signal peptide sequences and α-amylase inhibitor domain. There was no significant change in mutants' spatial conformation confirmed by Circular Dichroism. The position K132N and peptides at 108-117 and 132-141 were the core B- and T cell epitopes of Fag t 2 confirmed by competitive inhibition ELISA and dot blot. This result was of great significance on the study of allergen epitopes in prevention and treatment of hypersensitivity.


Assuntos
Alérgenos/imunologia , Epitopos/imunologia , Fagopyrum/metabolismo , Alérgenos/química , Alérgenos/genética , Alérgenos/metabolismo , Sequência de Aminoácidos , Dicroísmo Circular , Ensaio de Imunoadsorção Enzimática , Epitopos/genética , Epitopos/metabolismo , Mutagênese Sítio-Dirigida , Pichia/metabolismo , Proteínas Recombinantes/biossíntese , Proteínas Recombinantes/química , Proteínas Recombinantes/imunologia , Proteínas Recombinantes/isolamento & purificação
3.
Molecules ; 26(2)2021 Jan 06.
Artigo em Inglês | MEDLINE | ID: mdl-33419110

RESUMO

(1) Background: Non-specific lipid transfer proteins (nsLTPs), which belong to the prolamin superfamily, are potent allergens. While the biological role of LTPs is still not well understood, it is known that these proteins bind lipids. Allergen nsLTPs are characterized by significant stability and resistance to digestion. (2) Methods: nsLTPs from gold kiwifruit (Act c 10.0101) and pomegranate (Pun g 1.0101) were isolated from their natural sources and structurally characterized using X-ray crystallography (3) Results: Both proteins crystallized and their crystal structures were determined. The proteins have a very similar overall fold with characteristic compact, mainly α-helical structures. The C-terminal sequence of Act c 10.0101 was updated based on our structural and mass spectrometry analysis. Information on proteins' sequences and structures was used to estimate the risk of cross-reactive reactions between Act c 10.0101 or Pun g 1.0101 and other allergens from this family of proteins. (4) Conclusions: Structural studies indicate a conformational flexibility of allergens from the nsLTP family and suggest that immunoglobulin E binding to some surface regions of these allergens may depend on ligand binding. Both Act c 10.0101 and Pun g 1.0101 are likely to be involved in cross-reactive reactions involving other proteins from the nsLTP family.


Assuntos
Actinidia/química , Alérgenos/química , Antígenos de Plantas/química , Proteínas de Transporte/química , Proteínas de Plantas/química , Romã (Fruta)/química , Sementes/química , Alérgenos/isolamento & purificação , Antígenos de Plantas/isolamento & purificação , Proteínas de Transporte/isolamento & purificação , Cristalografia por Raios X , Proteínas de Plantas/isolamento & purificação , Conformação Proteica em alfa-Hélice
4.
J Agric Food Chem ; 69(4): 1379-1390, 2021 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-33464885

RESUMO

A high content of potentially allergenic lectin in Phaseolus vulgaris L. beans is of increasing health concerns; however, understanding of the protein allergenicity mechanism on the molecular basis is scarce. In the present study, low-pH treatments were applied to modify black turtle bean lectin allergen, and a sensitization procedure was performed using the BALB/c mice for the allergenicity evaluation, while the conformational changes were monitored by the spectral analyses and the details were explored by the molecular dynamics simulation. Much milder anaphylactic responses were observed in BALB/c mice experiments. At the molecular level, the protein was unfolded in low acidic environments because of protonation, and α-helix was reduced with the exposure of trypsin cleavage sites, especially the improvement of protease accessibility for Lys121, 134, and 157 in the B cell epitope structural alterations. These results indicate that a low-pH treatment might be an efficient method to improve the safety of legume protein consumption.


Assuntos
Alérgenos/química , Lectinas/química , Phaseolus/imunologia , Alérgenos/imunologia , Animais , Linfócitos B/imunologia , Feminino , Manipulação de Alimentos , Hipersensibilidade Alimentar/imunologia , Humanos , Concentração de Íons de Hidrogênio , Lectinas/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Phaseolus/genética , Sementes/química , Sementes/imunologia
5.
BMC Complement Med Ther ; 21(1): 41, 2021 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-33478471

RESUMO

BACKGROUND: The latest coronavirus SARS-CoV-2, discovered in China and rapidly spread Worldwide. COVID-19 affected millions of people and killed hundreds of thousands worldwide. There are many ongoing studies investigating drug(s) suitable for preventing and/or treating this pandemic; however, there are no specific drugs or vaccines available to treat or prevent SARS-CoV-2 as of today. METHODS: Fifty-eight fragrance materials, which are classified as allergen fragrance molecules, were selected and used in this study. Docking simulations were carried out using four functional proteins; the Covid19 Main Protase (MPro), Receptor binding domain (RBD) of spike protein, Nucleocapsid, and host Bromodomain protein (BRD2), as target macromolecules. Three different software, AutoDock, AutoDock Vina (Vina), and Molegro Virtual Docker (MVD), running a total of four different docking protocol with optimized energy functions were used. Results were compared with the five molecules reported in the literature as potential drugs against COVID-19. Virtual screening was carried out using Vina, molecules satisfying our cut-off (- 6.5 kcal/mol) binding affinity was confirmed by MVD. Selected molecules were analyzed using the flexible docking protocol of Vina and AutoDock default settings. RESULTS: Ten out of 58 allergen fragrance molecules were selected for further docking studies. MPro and BRD2 are potential targets for the tested allergen fragrance molecules, while RBD and Nucleocapsid showed weak binding energies. According to AutoDock results, three molecules, Benzyl Cinnamate, Dihydroambrettolide, and Galaxolide, had good binding affinities to BRD2. While Dihydroambrettolide and Galaxolide showed the potential to bind to MPro, Sclareol and Vertofix had the best calculated binding affinities to this target. When the flexible docking results analyzed, all the molecules tested had better calculated binding affinities as expected. Benzyl Benzoate and Benzyl Salicylate showed good binding affinities to BRD2. In the case of MPro, Sclareol had the lowest binding affinity among all the tested allergen fragrance molecules. CONCLUSION: Allergen fragrance molecules are readily available, cost-efficient, and shown to be safe for human use. Results showed that several of these molecules had comparable binding affinities as the potential drug molecules reported in the literature to target proteins. Thus, these allergen molecules at correct doses could have significant health benefits.


Assuntos
Alérgenos/química , Alérgenos/imunologia , /imunologia , Odorantes , Perfumes/química , /imunologia , Monofosfato de Adenosina/análogos & derivados , Monofosfato de Adenosina/química , Monofosfato de Adenosina/metabolismo , Alanina/análogos & derivados , Alanina/química , Alanina/metabolismo , Alérgenos/administração & dosagem , Alérgenos/uso terapêutico , Benzopiranos/química , Benzopiranos/metabolismo , Compostos de Benzil/química , Compostos de Benzil/metabolismo , Cinamatos/química , Cinamatos/metabolismo , /metabolismo , /metabolismo , Diterpenos/química , Diterpenos/metabolismo , Avaliação Pré-Clínica de Medicamentos , Humanos , Ligantes , Simulação de Acoplamento Molecular , Perfumes/administração & dosagem , Perfumes/uso terapêutico , Fosfoproteínas/química , Fosfoproteínas/metabolismo , Glicoproteína da Espícula de Coronavírus/química , Glicoproteína da Espícula de Coronavírus/metabolismo , Fatores de Transcrição/química , Fatores de Transcrição/metabolismo
6.
Food Chem ; 339: 128106, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-33152886

RESUMO

It is practical to inhibit the allergenicity of ß-lactoglobulin (ß-LG) using natural products acting via noncovalent interactions; however, the mechanism of the effect has not been investigated in detail. Herein, the comprehensive noncovalent mechanism of inhibition of the antigenicity of ß-LG by six flavonoids (kaempferol, myricetin, phloretin, epigallocatechin-3-gallate (EGCG), naringenin, and quercetin) was investigated by spectroscopic and molecular docking methods. Our results indicate that six flavonoids reduced the antigenicity of ß-LG in the following order: EGCG > phloretin > naringenin > myricetin > kaempferol > quercetin, with antigenic inhibition rates of 72.6%, 68.4%, 59.7%, 52.3%, 51.4% and 40.8%, respectively. Six flavonoids induced distinct conformational changes in ß-LG, which were closely associated with a decline in antigenicity of ß-LG. The flavonoids bound to specific antigen epitopes in the ß-sheet and ß-turn of ß-LG to induce a decrease in the antigenicity of the protein.


Assuntos
Flavonoides/farmacologia , Lactoglobulinas/química , Lactoglobulinas/imunologia , Alérgenos/química , Alérgenos/imunologia , Alérgenos/metabolismo , Dicroísmo Circular , Epitopos/metabolismo , Flavonoides/química , Flavonoides/metabolismo , Lactoglobulinas/metabolismo , Simulação de Acoplamento Molecular , Conformação Proteica , Espectrometria de Fluorescência , Espectroscopia de Infravermelho com Transformada de Fourier , Análise Espectral
7.
Food Chem ; 339: 127895, 2021 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-32866706

RESUMO

The epitopes of the major allergen of pine nut, Pin p 1, were analyzed using a peptide library and sera from patients with clinical allergy to pine nut in order to deepen into the allergenic characteristics of Pin p 1. Analyses of epitope similarities and epitopes location in a 3D-model were also performed. Results showed that three main regions of Pin p 1 containing 5 epitopes were recognized by patient sera IgE. The epitopes of Pin p 1 had important similarities with epitopes of allergenic 2S albumins from peanut (Ara h 2 and 6) and Brazil nut (Ber e 1). The epitopes of Pin p 1 were found in α-helices and coils in the 3D protein structure. Interestingly, all epitopes were found to be well-exposed in the protein surface, which suggests facile access for IgE-binding to the structure of Pin p 1 which is known to be highly resistant.


Assuntos
Albuminas 2S de Plantas/química , Alérgenos/química , Mapeamento de Epitopos/métodos , Epitopos/química , Pinus/metabolismo , Albuminas 2S de Plantas/imunologia , Albuminas 2S de Plantas/metabolismo , Adolescente , Adulto , Alérgenos/imunologia , Sequência de Aminoácidos , Arachis/imunologia , Arachis/metabolismo , Epitopos/imunologia , Feminino , Humanos , Imunoglobulina E/sangue , Imunoglobulina E/imunologia , Masculino , Hipersensibilidade a Noz/imunologia , Hipersensibilidade a Noz/patologia , Nozes/imunologia , Nozes/metabolismo , Biblioteca de Peptídeos , Pinus/imunologia
8.
Food Chem ; 337: 127986, 2021 Feb 01.
Artigo em Inglês | MEDLINE | ID: mdl-32920269

RESUMO

We have developed a novel approach that involves inception-resnet network (IRN) modeling based on infrared spectroscopy (IR) for rapid and specific detection of the fish allergen parvalbumin. SDS-PAGE and ELISA were used to validate the new method. Through training and learning with parvalbumin IR spectra from 16 fish species, IRN, support vector machine (SVM), and random forest (RF) models were successfully established and compared. The IRN model extracted highly representative features from the IR spectra, leading to high accuracy in recognizing parvalbumin (up to 97.3%) in a variety of seafood matrices. The proposed infrared spectroscopic IRN (IR-IRN) method was rapid (~20 min, cf. ELISA ~4 h) and required minimal expert knowledge for application. Thus, it could be extended for large-scale field screening and identification of parvalbumin or other potential allergens in complex food matrices.


Assuntos
Produtos Pesqueiros/análise , Proteínas de Peixes/análise , Redes Neurais de Computação , Parvalbuminas/análise , Espectrofotometria Infravermelho/estatística & dados numéricos , Alérgenos/química , Animais , Eletroforese em Gel de Poliacrilamida , Ensaio de Imunoadsorção Enzimática , Peixes/imunologia , Análise de Alimentos/métodos , Análise de Alimentos/estatística & dados numéricos , Hipersensibilidade Alimentar , Camundongos Endogâmicos BALB C , Parvalbuminas/imunologia , Reprodutibilidade dos Testes , Espectrofotometria Infravermelho/métodos , Máquina de Vetores de Suporte
9.
Methods Mol Biol ; 2178: 357-376, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33128761

RESUMO

Matrix-assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is largely recognized as an important tool in the analysis of many biomolecules such as proteins and peptides. The MS analysis of digested peptides to identify a protein or some of its modifications is a key step in proteomics. MALDI-MS is well suited for the peptide mass fingerprinting (PMF) technique, as well as selected fragmentation of various precursors using collisional-induced dissociation (CID) or post-source decay (PSD).In the last few years, MALDI-MS has played a significant role in food chemistry, especially in the detection of food adulterations, characterization of food allergens, and investigation of protein structural modifications induced by various industrial processes that could be an issue in terms of food quality and safety.Here, we present simple extraction protocols of allergenic proteins in food commodities such as milk, egg, hazelnut , and lupin seeds. Classic bottom-up approaches based on Sodium Dodecyl Sulphate (SDS) gel electrophoresis separation followed by in-gel digestion or direct in-solution digestion of whole samples are described. MALDI-MS and MS /MS analyses are discussed along with a comparison of data obtained by using the most widespread matrices for proteomic studies, namely, α-cyano-4-hydroxy-cinnamic acid (CHCA) and α-cyano-4-chloro-cinnamic acid (CClCA). The choice of the most suitable MALDI matrix is fundamental for high-throughput screening of putative food allergens.


Assuntos
Alérgenos/análise , Análise de Alimentos , Hipersensibilidade Alimentar , Proteômica , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Alérgenos/química
10.
Methods Mol Biol ; 2223: 1-17, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33226583

RESUMO

Mouse models of allergic disease offer numerous advantages when compared to the models of other animals. However, selection of appropriate mouse models is critical to advance the field of food allergy by revealing mechanisms of allergy and for testing novel therapeutic approaches. All current mouse models for food allergy have weaknesses that may limit their applicability to human disease. Aspects such as the genetic predisposition to allergy or tolerance from the strain of mouse used, allergen dose, route of exposure (oral, intranasal, intraperitoneal, or epicutaneous), damage of the epithelial barrier, use of adjuvants, food matrix effects, or composition of the microbiota should be considered prior to the selection of a specific murine model and contemplated according to the intended purpose of the study. This chapter reviews our current knowledge on the application of mouse models to food allergy research and the variables that may influence the successful development of each type of model.


Assuntos
Adjuvantes Imunológicos/administração & dosagem , Alérgenos/administração & dosagem , Modelos Animais de Doenças , Hipersensibilidade Alimentar/imunologia , Regulação da Expressão Gênica/imunologia , Alérgenos/química , Animais , Toxinas Bacterianas/administração & dosagem , Misturas Complexas/administração & dosagem , Misturas Complexas/química , Vias de Administração de Medicamentos , Hipersensibilidade Alimentar/etiologia , Hipersensibilidade Alimentar/genética , Hipersensibilidade Alimentar/patologia , Fatores de Troca do Nucleotídeo Guanina/genética , Fatores de Troca do Nucleotídeo Guanina/imunologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C3H , Camundongos Knockout , Receptores de Superfície Celular/genética , Receptores de Superfície Celular/imunologia , Especificidade da Espécie , Receptor 4 Toll-Like/genética , Receptor 4 Toll-Like/imunologia
11.
PLoS One ; 15(6): e0233745, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32542029

RESUMO

The susceptibility of newly expressed proteins to digestion by gastrointestinal proteases (e.g., pepsin) has long been regarded as one of the important endpoints in the weight-of-evidence (WOE) approach to assess the allergenic risk of genetically modified (GM) crops. The European Food Safety Authority (EFSA) has suggested that current digestion study protocols used for this assessment should be modified to more accurately reflect the diverse physiological conditions encountered in human populations and that the post-digestion analysis should include analytical methods to detect small peptide digestion products.The susceptibility of two allergens (beta-lactoglobin (ß-Lg) and alpha-lactalbumin (α-La)) and two non-allergens (hemoglobin (Hb) and phosphofructokinase (PFK)) to proteolytic degradation was investigated under two pepsin digestion conditions (optimal pepsin digestion condition: pH 1.2, 10 U pepsin/µg test protein; sub-optimal pepsin digestion condition: pH 5.0, 1 U pepsin/10 mg test protein), followed by 34.5 U trypsin/mg test protein and 0.4 U chymotrypsin/mg test protein digestion in the absence or presence of bile salts. All samples were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) in conjunction with Coomassie Blue staining and, in parallel, liquid chromatography tandem mass spectrometry (LC-MS) detection. The results provide following insights: 1) LC-MS methodology does provide the detection of small peptides; 2) Peptides are detected in both allergens and non-allergens from all digestion conditions; 3) No clear differences among the peptides detected from allergen and non-allergens; 4) The differences observed in SDS-PAGE between the optimal and sub-optimal pepsin digestion conditions are expected and align with kinetics and properties of the specific enzymes; 5) The new methodology with new digestion conditions and LC-MS detection does not provide any differentiating information for prediction whether a protein is an allergen. The classic pepsin resistance assay remains the most useful assessment of the potential exposure of an intact newly expressed protein as part of product safety assessment within a WOE approach.


Assuntos
Alérgenos/química , Análise de Alimentos/métodos , Peptídeos/química , Proteólise , Alérgenos/metabolismo , Animais , Cromatografia Líquida/métodos , Inocuidade dos Alimentos , Hemoglobinas/química , Hemoglobinas/metabolismo , Lactalbumina/química , Lactalbumina/metabolismo , Lactoglobulinas/química , Lactoglobulinas/metabolismo , Peptídeos/metabolismo , Fosfofrutoquinases/química , Fosfofrutoquinases/metabolismo , Suínos , Espectrometria de Massas em Tandem/métodos , Tripsina/metabolismo
12.
Food Chem ; 326: 127027, 2020 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-32438232

RESUMO

This work reports on theeffect of heat treatment on the protein conformational stabilityof intact and post-translationallycleaved peanut allergen Ara h 6 in relation to IgE-binding. Intact and post-translationallycleaved Ara h 6 are structurally similar and theirstrong resistance to denaturant-inducedunfolding is comparable. Only upon exposure toautoclave conditions the twoforms of Ara h 6 demonstrated susceptibility toirreversible denaturationresulting in a significant decrease in IgE-binding potency. Thisreduction isfor the intact protein more pronounced than for than for the cleaved form. This isattributed to less conformational constrains of the cleaved form comparedtointact, as suggested by the 2-fold lower activation energy for unfoldingfound for the cleavedform. Overall, harsh conditionsare required to denature Ara h 6 and to significantly reduce its IgE-bindingpotency. The cleavedform possesses more resistance to such denaturation than the intactform.


Assuntos
Albuminas 2S de Plantas/química , Alérgenos/química , Antígenos de Plantas/química , Arachis/química , Imunoglobulina E/imunologia , Albuminas 2S de Plantas/imunologia , Alérgenos/imunologia , Antígenos de Plantas/imunologia , Temperatura Alta , Conformação Proteica , Fatores de Tempo
13.
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
14.
J Dairy Sci ; 103(5): 4109-4120, 2020 May.
Artigo em Inglês | MEDLINE | ID: mdl-32113777

RESUMO

Bovine ß-lactoglobulin (ß-LG) is recognized as a major allergen in milk. This study aimed to investigate ultrasound-assisted irradiation for reducing the allergenicity of ß-LG, since irradiation can reduce the allergenicity of cow milk proteins and ultrasound can improve the quality of milk. The structural changes induced in high purity ß-LG, treated by irradiation, with or without sonication, were characterized by native PAGE, circular dichroism spectroscopy, and fluorescence spectroscopy. The changes in allergenicity were measured by IgE binding capacity to, and inflammatory mediator secretion by, human basophil KU812 cells. Surface hydrophobicity was reduced and aggregation of ß-LG increased after treatment by irradiation, both with and without sonication. The IgE binding capacity and release of inflammatory mediators were reduced significantly and the reduction induced by irradiation before sonication was the greatest, suggesting that irradiation after sonication can be a safe and effective method to reduce the allergenicity of ß-LG in dairy processing.


Assuntos
Alérgenos/química , Lactoglobulinas/química , Lactoglobulinas/imunologia , Leite/química , Alérgenos/imunologia , Animais , Bovinos , Pré-Escolar , Dicroísmo Circular , Eletroforese em Gel de Poliacrilamida , Feminino , Humanos , Interações Hidrofóbicas e Hidrofílicas , Imunoglobulina E/imunologia , Lactente , Lactoglobulinas/efeitos da radiação , Masculino , Hipersensibilidade a Leite , Conformação Proteica , Sonicação , Espectrometria de Fluorescência
15.
J Food Sci ; 85(3): 789-799, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-32078753

RESUMO

To investigate the influence of heat treatment and egg matrix on egg custard (EC) proteins, 12 different kinds of ECs with different egg/water ratios (1:1, 1:1.5, 1:2, or 1:3, v/v) and different heating temperatures (80, 90, or 100 °C) and times (10, 15, or 20 min) were prepared and evaluated for the digestibility, structure, eliciting capacity and sensitizing capacity using SDS-PAGE, fluorescence spectra, ELISA, and a BALB/c mouse model, respectively. The physicochemical properties of EC proteins were significantly affected by heat treatment and egg matrix, which showed the increased digestibility and partially unfolded structure. The eliciting capacity of EC evaluated by IgE binding to sera from egg-allergic patients was reduced after heat treatment, and the EC made by heating at 100 °C for 20 min with a whole egg/water ratio of 1:2 (v/v) was the weakest. The sensitizing capacity of EC was also reduced in the BALB/c mouse model, which showed the significantly decreased levels of specific IgE, IgG, IgG1 and IgG2a, mMCP-1 and histamine in the mouse sera, as well as cytokine secretions of IL-4, IL-5, and IL-13, compared with the raw egg (RE) group. Results demonstrate that heat treatment and egg matrix significantly reduced the eliciting and sensitizing capacity of EC by changing the tertiary structure and increasing the digestibility of EC proteins. PRACTICAL APPLICATION: Egg custard (EC) is one kind of savory food suitable for all ages, and is also a traditional supplementary food for infants and young children in China. However, limited information is available on the allergenicity of egg custard. In this work, we evaluated how the structure, digestibility, and allergenic potential of egg allergens in EC were altered by the degree of thermal treatment and egg matrix, and elucidated the links between the physicochemical properties and allergenic potential of EC affected by heat treatment and egg matrix. Our results demonstrate that heat treatment and egg matrix significantly reduced the eliciting and sensitizing capacity of EC by changing the tertiary structure and increasing the digestibility of EC proteins.


Assuntos
Alérgenos/química , Alérgenos/imunologia , Culinária/métodos , Hipersensibilidade a Ovo/imunologia , Proteínas do Ovo/química , Proteínas do Ovo/imunologia , Animais , Galinhas , Criança , Pré-Escolar , China , Hipersensibilidade a Ovo/sangue , Hipersensibilidade a Ovo/prevenção & controle , Ensaio de Imunoadsorção Enzimática , Feminino , Manipulação de Alimentos , Temperatura Alta , Humanos , Imunoglobulina E/sangue , Imunoglobulina G/sangue , Lactente , Masculino , Camundongos , Camundongos Endogâmicos BALB C
16.
Mol Immunol ; 120: 1-12, 2020 04.
Artigo em Inglês | MEDLINE | ID: mdl-32044430

RESUMO

Mus m 1.0102 is a member of the mouse Major Urinary Protein family, belonging to the Lipocalins superfamily. Major Urinary Proteins (MUPs) are characterized by highly conserved structural motifs. These include a disulphide bond, involved in protein oxidative folding and protein structure stabilization, and a free cysteine residue, substituted by serine only in the pheromonal protein Darcin (MUP20). The free cysteine is recognized as responsible for the onset of inter- or intramolecular thiol/disulphide exchange, an event that favours protein aggregation. Here we show that the substitution of selected cysteine residues modulates Mus m 1.0102 protein folding, fold stability and unfolding reversibility, while maintaining its allergenic potency. Recombinant allergens used for immunotherapy or employed in allergy diagnostic kits require, as essential features, conformational stability, sample homogeneity and proper immunogenicity. In this perspective, recombinant Mus m 1.0102 might appear reasonably adequate as lead molecule because of its allergenic potential and thermal stability. However, its modest resistance to aggregation renders the protein unsuitable for pharmacological preparations. Point mutation is considered a winning strategy. We report that, among the tested mutants, C138A mutant acquires a structure more resistant to thermal stress and less prone to aggregation, two events that act positively on the protein shelf life. Those features make that MUP variant an attractive lead molecule for the development of a diagnostic kit and/or a vaccine.


Assuntos
Alérgenos/química , Alérgenos/imunologia , Proteínas/química , Proteínas/imunologia , Alérgenos/genética , Substituição de Aminoácidos , Animais , Linhagem Celular , Cisteína/química , Humanos , Testes Imunológicos , Ligantes , Camundongos , Modelos Moleculares , Mutagênese Sítio-Dirigida , Conformação Proteica , Dobramento de Proteína , Estabilidade Proteica , Estrutura Secundária de Proteína , Proteínas/genética , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Espectrometria de Fluorescência , Espectroscopia de Infravermelho com Transformada de Fourier
17.
Molecules ; 25(4)2020 Feb 20.
Artigo em Inglês | MEDLINE | ID: mdl-32093394

RESUMO

Tree nuts confer many health benefits due to their high content of vitamins and antioxidants, and they are increasingly consumed in the last few years. Food processing is an important industrial tool to modify allergenic properties of foods, in addition to ensuring safety and enhancing organoleptic characteristics. The effect of high pressure, without and with heating, on SDS-PAGE and immunodetection profile of potential allergenic proteins (anti-11S, anti-2S and anti-LTP) of pistachio, cashew, peanut, hazelnut, almond, and chestnut was investigated. Processing based on heat and/or pressure and ultra-high pressure (HHP, 300-600 MPa) without heating was applied. After treating the six tree nuts with pressure combined with heat, a progressive diminution of proteins with potential allergenic properties was observed. Moreover, some tree nuts proteins (pistachio, cashew, and peanut) seemed to be more resistant to technological processing than others (hazelnut and chestnut). High pressure combined with heating processing markedly reduce tree nut allergenic potential as the pressure and treatment time increases. HHP do not alter hazelnut and almond immunoreactivity.


Assuntos
Alérgenos/química , Manipulação de Alimentos , Nozes/química , Proteínas de Plantas/química , Alérgenos/imunologia , Hipersensibilidade Alimentar , Temperatura Alta , Humanos , Nozes/imunologia , Proteínas de Plantas/imunologia , Pressão
18.
Allergol. immunopatol ; 48(1): 26-33, ene.-feb. 2020. ilus, graf
Artigo em Inglês | IBECS | ID: ibc-186588

RESUMO

Introduction and Objectives: The production and consumption of oysters is increasing annually because it can provide essential nutrients and benefit for human health, leading to frequent occurrence of severe allergic reactions observed in sensitized individuals. The aim of the present study was to investigate the effects of acid and protease treatment on the conformation and IgE-binding capacity of recombinant Crassostrea gigas tropomyosin (Cra g 1). Results: Under acidic conditions, Cra g 1 did not undergo degradation, however, the changes obvious in the intensity of CD signal and ANS-binding fluorescence were observed, which was associated with a decrease in antibody reactivity. In simulated gastrointestinal fluid (SGF) and simulated intestinal fluid (SIF) digestion system, acid-treated Cra g 1 was relatively resistant to digestion, but the degradative patterns were very different. Moreover, owing to alterations of secondary structure and hydrophobic surface of the protein during digestive processing, antigenicity of acid-induced Cra g 1 reduced in SGF while it increased significantly in SIF. Conclusion: To our knowledge, this is the first study reporting that antigenicity of acid-treated oyster tropomyosin increased after SIF digestion. These results revealed that treatment with acid and pepsin, rather than trypsin, was an effective way of reducing IgE-binding capacity of tropomyosin from oyster


No disponible


Assuntos
Humanos , Imunoglobulina E/imunologia , Técnicas In Vitro/métodos , Alérgenos/química , Tropomiosina/química , Hipersensibilidade Alimentar/imunologia , Digestão , Alérgenos/imunologia , Tropomiosina/imunologia , Ostreidae/imunologia , Sistema Digestório/imunologia , Eletroforese/métodos , Análise Espectral/métodos , Ensaio de Imunoadsorção Enzimática
19.
Molecules ; 25(2)2020 Jan 16.
Artigo em Inglês | MEDLINE | ID: mdl-31963206

RESUMO

Four recombinant (r) allergens (rAmb a 8.0101, rArt v 4.0101, rBet v 2.0101, and rPhl p 12.0101) were successfully produced and used for sensitization studies. The allergens belong to the profilin family which is one of the most numerous allergen families. These four proteins represent allergens originating from pollen of weeds (rAmb a 8.0101 and rArt v 4.0101), tree (rBet v 2.0101) and grass (rPhl p 12.0101). The recombinant allergens were characterized using various biochemical and biophysical methods and tested for their ability to bind patient-derived antibodies. One hundred patients aged 2 to 50 years sensitized to pollen and plant-derived food allergens (IgE > 0.35 kU/L) were included. Sensitization to individual allergen sources and components of birch and timothy pollens was evaluated using multiparameter immunoblots. The presence of IgE to pollen-derived recombinant profilins rAmb a 8.0101, rArt v 4.0101, rBet v 2.0101, and rPhl p 12.0101 in serum was evaluated using ELISA method. The presence of IgE against pollen profilins was detected in 20 out of 100 studied patients. High correlation was seen between IgE ELISA results with individual pollen profilins. In summary, it was shown that the recombinant versions of the four allergenic profilins can be used for sensitization studies and for component-resolved allergy diagnostics.


Assuntos
Alérgenos/imunologia , Antígenos de Plantas/imunologia , Hipersensibilidade/imunologia , Profilinas/imunologia , Proteínas Recombinantes/imunologia , Alérgenos/química , Sequência de Aminoácidos , Antígenos de Plantas/química , Imunização , Modelos Moleculares , Profilinas/química , Conformação Proteica , Estabilidade Proteica , Proteínas Recombinantes/química , Análise Espectral , Relação Estrutura-Atividade , Termodinâmica
20.
Food Chem ; 313: 126019, 2020 May 30.
Artigo em Inglês | MEDLINE | ID: mdl-31931421

RESUMO

Recalls of spice containing products due to undeclared peanut have highlighted the importance of analytical methods in these foods. We examined the performance of peanut detection methods in cumin and garlic, focusing on quantitative ELISA. Although suitable for qualitative detection, accurate quantitation proved difficult. Roasting of peanut contaminants influenced ELISA results, with raw peanut over-detected (3.9-fold) and roasted peanut under-detected (3.5-fold). Further investigation demonstrated the importance of protein targets for ELISA. The kit which gave the least variable results was based on detection of 2S albumin proteins. Additionally, we show that these proteins are more efficiently extracted from roasted peanut. We conclude that current methods are largely suitable for the qualitative detection of peanut in cumin and garlic. Quantitation relies on assumptions as to the state of thermal processing of peanut. We suggest that analytical method providers address robust detection by target selection, including identifying targets by MS.


Assuntos
Alérgenos/análise , Arachis/química , Alérgenos/química , Sequência de Aminoácidos , Arachis/metabolismo , Cromatografia Líquida de Alta Pressão , Cuminum/química , Ensaio de Imunoadsorção Enzimática , Temperatura Alta , Espectrometria de Massas , Peptídeos/análise , Peptídeos/química , Proteínas de Plantas/análise
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA
...