Allergenic Proteins Research Articles

Occupational Rhinoconjunctivitis Induced by Flaxseed: Identification of Novel Allergenic Proteins From Cupin, Chaperonin, and Enzyme Families.

Occupational Rhinoconjunctivitis Induced by Flaxseed: Identification of Novel Allergenic Proteins From Cupin, Chaperonin, and Enzyme Families.

Analysis of the sensitization activity of Moringa oleifera leaves protein

The determination of allergenic proteins in Moringa oleifera leaves, which is the main components of immune activity, has enabled the development of a more effective method for evaluating the activity of extracted Moringa oleifera leaves protein. In this study, the extraction process of Moringa oleifera leaves protein was optimized based on a single factor experiment. The hemagglutination-related properties of Moringa oleifera leaves protein, such as (thermal, acid–base) stability, sugar binding specificity, ion binding characteristics, and hemolytic activity, were detected. The optimal combination of extraction process was: extraction time of 6 h, material-liquid ratio of 1:8, and ammonium sulfate saturation of 60%. The extraction rate of moringa leaf protein under this condition was 14.37 mg/g. The molecular weight of moringa leaf protein was analyzed by SDS-PAGE, and the molecular weight was mainly concentrated around 23 kDa~70 kDa, with the highest content of 35 kDa (major allergen). The study of the hemagglutination characteristics of Moringa oleifera leaves protein revealed that the protein exhibited high stability at temperatures below 60°C, with complete loss of activity occurring at temperatures above 110°C for 20 min. The effect of different pH conditions on the hemagglutination capacity of Moringa oleifera leaves protein was readily discernible. The hemagglutination activity of Moringa oleifera leaves protein was 104 in a pH value from 3.7 to 7.8, and the hemagglutination activity was completely lost at a pH value higher than 11.9. D(+) anhydrous glucose is the specific inhibitory sugar of Moringa oleifera leaves protein lectin. Moringa oleifera leaves protein exhibits hemolytic activity at a concentration of at least 20 mg/mL, and α-methyl-mannoside, galactoside, raffinose and Al3+ can inhibit the hemolysis of Moringa oleifera leaves protein. The present study identified the effects of different factors on the coagulation activity and hemolytic ability of Moringa oleifera leaves protein, thereby providing a theoretical basis for further purification and application of Moringa oleifera lectin. However, it should be noted that the results of the mixture have certain limitations, and further purification of lectin is needed to obtain more targeted research results.

Randomized, Placebo-Controlled, Phase 1 Safety Study of Oral Mucosal Immunotherapy in Peanut Allergic Adults.

Oral Mucosal Immunotherapy (OMIT) uses a specifically formulated toothpaste to deliver allergenic proteins to immunologically active areas of the oral cavity. This represents a new delivery mechanism with several features designed to improve food allergy desensitization. OMIT presents advantages over other approaches to allergy immunotherapy due to its targeted delivery and simplified administration. To determine the safety, tolerability, and adherence of OMIT with INT301 in peanut allergic adults. This Oral Mucosal Escalation Goal Assessment (OMEGA) study enrolled 32 adults, age 18-55, with peanut allergy in a 3:1 ratio to receive either an escalating dose of INT301 or placebo. Entry criteria included a positive SPT with a wheal diameter ≥ 3mm greater than control and/or psIgE ≥ 0.35 kU/L. Subjects were required to fail an oral food challenge ≤ 100 mg dose of peanut protein. Safety and tolerability were monitored during this 48-week trial. 100% of active subjects tolerated the pre-specified protocol highest dose. No moderate nor severe systemic reactions occurred in active participants. Non-systemic adverse reactions were mostly local (oral and nasal cavity), mild and transient. Active subjects adhered to treatment 97% of days on study, with no withdrawals due to study medication. In the OMEGA trial, INT301 met all primary and secondary endpoints of safety, tolerability, and adherence. OMIT appears to be a safe and convenient option for individuals with food allergies. These results support its further evaluation in the pediatric population.

Dual-protein system composed of soy protein and β-lactoglobulin: effect of transglutaminase-mediated crosslinking on its allergenicity and conformation.

The escalating global prevalence of food allergies has intensified the need for hypoallergenic food products. Transglutaminase (TGase)-mediated crosslinking has garnered significant attention for its potential to reduce the allergenicity of food proteins. This study aimed to investigate the effects of TGase crosslinking on the potential allergenicity and conformational changes in a dual-protein system composed of β-lactoglobulin (β-LG) and soy protein isolate (SPI) at varying mass ratios (10:0, 7:3, 5:5, 3:7 and 0:10 (w/w)). TGase preferentially crosslinked the 7S and 11S subunits of soy protein, rather than β-LG. Crosslinking treatment reduced the allergenic potential of both soy protein and β-LG, with the degree of reduction depending on the protein ratio. The β-LG5:SPI5 and β-LG7:SPI3 mixtures showed the most significant reduction in antibody reactivity towards soy protein and β-LG, respectively. Additionally, TGase-mediated crosslinking significantly reduced the binding capacity of all dual-protein samples to serum immunoglobulin E from allergic patients, compared to the control group (P < 0.05). The allergenicity reduction was accompanied by structural modifications, including a decrease in β-sheet content, an increase in β-turn and random coil structures, enhanced ultraviolet absorption and intrinsic fluorescence, reduced free sulfhydryl levels and altered intermolecular forces. These changes suggest that TGase-induced crosslinking may disrupt or mask allergenic epitopes, thus lowering allergenicity. TGase crosslinking effectively reduced the allergenic potential of the dual-protein system by inducing structural changes. These findings underscore the potential of TGase in developing hypoallergenic dual-protein food products and provide a scientific foundation for future research and application in the food industry. © 2025 Society of Chemical Industry.

Edible Insects as an Alternative Source of Nutrients: Benefits, Risks, and the Future of Entomophagy in Europe-A Narrative Review.

According to projections by the Food and Agriculture Organization of the United Nations, the global population will reach 9 billion by 2050. This raises concerns about the ability to feed such a population. In view of the above, it is necessary to search for alternative food sources. Edible insects are rich in complete protein, essential fatty acids, vitamins and micronutrients. Despite this, entomophagy is not common in Europe. In 2021, the European Union approved Acheta domesticus, Tenebrio molitor, Locusta migratoria, and Alphitobius diaperinus for consumption. However, their consumption may also be associated with certain hazards, e.g., food allergies. The purpose of this review is to present existing knowledge, discuss the possible dangers of consuming insects, and identify areas for further research. Studies in Asian populations indicate that edible insects may be responsible for 4.2-19.4% of food allergies and 18% of fatal food-induced anaphylaxis. There are also increasing reports from Europe of food allergies to edible insects. A thorough understanding of allergens, their properties, and the mechanisms of food allergies associated with edible insects' consumption is essential for ensuring consumers' safety. In the future, it would be worthwhile to investigate the effects of heat treatment on the allergenicity of insect proteins.

Structural determinants of peanut induced anaphylaxis.

Human monoclonal IgE antibodies recognizing peanut allergens have recently become available, but we lack a detailed understanding of how these IgEs target allergens. To determine the molecular details of the antibody-allergen interaction for a panel of clinically important human IgE monoclonal antibodies and to develop strategies to disrupt disease causing antibody-allergen interactions. We identified candidates from a panel of epitope binned human IgE monoclonals that recognize two important and homologous peanut allergens, Ara h 2 and Ara h 6. Crystal structures were determined revealing the interfaces (antigenic sites) of exemplars of five common IgE bins. Among the common antigenic sites on Ara h 2 and Ara h 6, two sites (A and B) are highly conserved between the allergens, explaining the cross reactivity of antibodies that recognize these sites. Three sites (C, D, and F) involve residues that are not conserved between the allergens. Of the five common sites, three Sites (B, C, and D) involve residues that are only near each other when the allergens are properly folded, such that these sites are conformational. Two additional sites (sites A and F) involve largely linear stretches of amino acids. Site F targeted antibody, 38B7, binds to a peptide sequence DPYSPOHS, in which hydroxylation of the last proline is critical for binding. This sequence is repeated two or three times depending on the Ara h 2 isoform, enabling 38B7 to induce anaphylaxis as a single monoclonal, without a second antibody. We have mutated key residues in each site and created a panel of hypoallergens, having reduced IgE mAb binding and lacking the ability to induce anaphylaxis in our murine model. We created a structural map of the IgE antibody response to the most important peanut allergen proteins to enable the design of new allergy immunotherapies and vaccines.

Dietary Linolenic Acid Enhances IgE Binding to Bovine α-Lactalbumin/β-Lactoglobulin and Promotes KU812 Basophil Degranulation via Upregulation of the Lyn/Syk Pathway.

Milk proteins possess an abundance of free amino groups and exhibit diverse spatial structures. During food processing, these properties facilitate their interaction with hydrophobic ligands, such as linolenic acid. Exploring the IgE and IgG binding ability of linolenic acid-milk protein complexes at different temperatures, times, and molar ratios is crucial for controlling the allergenicity of milk proteins in food processing. In this study, the results indicate that linolenic acid can enhance the allergenicity of milk proteins. Moreover, by studying Lyn, Syk, NF-κB, and MAPK family-related proteins in the IgE/FcεRI-mediated signaling pathway, it is found that linolenic acid enhances cow's milk protein sensitization through the Lyn/Syk pathway. Our findings provide a further understanding of the interaction between milk nutrients and milk protein allergenicity.

Immunoproteomic analysis and identification of possible allergenic proteins in Artemisia annua pollen.

Artemisia annua (A. annua) is a wind-pollinated weed and a major allergen responsible for allergic respiratory diseases in Northern China. This study involved the separation of pollen proteins from A. annua utilizing SDS-PAGE and Coomassie Blue staining techniques. The effectiveness of extracting allergens from Artemisia annua pollen (AAP) were confirmed both in vivo and in vitro. A mouse model was established using A. annua pollen (AAP) extracts. In vitro, the interaction between allergenic proteins in the AAP extract and specific IgE antibodies present in patients' sera was analyzed, using IgE-immunoblotting and ELISA methods. Protein bands were subsequently analyzed by mass spectrometry. The recombinant Art an 3 (rArt an 3) protein was obtained via recombination, expression, and purification. Finally, the binding activity of rArt an 3 specific IgE was subsequently examined using Western blot and inhibition ELISA (iELISA). The electrophoretic profiles of AAP showed band patterns ranging from 10 to 70kDa, with the most prominet IgE-binding pollen proteins detected at approximately 12kDa. After stimulation of AAP, the sensitized group of mice exhibited significant allergic symptoms compared to the control group. Mass spectrometry analysis identified 7 proteins, including putative aldehyde oxidase Art an 7, Art v 1-like protein, Art an 3.0101 allergen precursor, Art an 3.0102 allergen precursor, Art an 3.0201 allergen precursor, lipid transfer protein 3, and non-specific lipid-transfer protein. The results of immunoblotting and iELISA showed that rArt an 3 could bind to IgE antibodies in the patient sera, and co-incubation of rArt an 3 with serum, the binding serum significantly inhibited IgE binding to the crude AAP. The Art an 3 is a key allergenic protein in AAP with a high IgE sensitization rate in the studied population sample. These findings enhance our understanding of the sensitization components of AAP and its sensitization characteristics within the Chinese population, thereby promoting the development of precise molecular diagnostics and therapeutic interventions.

Antigenic determinants underlying IgE-mediated anaphylaxis to peanut.

Studies of human IgE and its targeted epitopes on allergens have been very limited. We have an established method to immortalize IgE encoding B cells from allergic individuals. To develop an unbiased and comprehensive panel of peanut-specific human IgE mAbs to characterize key immunodominant antigenic regions and epitopes on peanut allergens to map the molecular interactions responsible for inducing anaphylaxis. Using human hybridoma technology to immortalize IgE encoding B cells from peripheral blood of subjects with severe peanut allergy, we generated a panel of naturally occurring human IgE mAbs in an unbiased manner. Isolated IgE mAbs were characterized extensively in allergen binding assays, peptide array analysis, antigenic mapping, binding kinetic analysis, serum blocking, skin testing inhibition, and functional assessment using human FcεRI transgenic mice. We created a large panel of 54 peanut-specific IgE mAbs, of which 63% were specific for Ara h 2 and/or 6. Pairs of IgE mAbs with the same antigen-specificity but different binding sites were able to mediate passive systemic anaphylaxis in FcεRI transgenic mice. A single mAb targeting the repetitive motif on Ara h 2 was able to induce degranulation and anaphylaxis on its own. IgG1 switched-variant immunoglobulins of the IgE mAb inhibited patients´ IgE binding to peanut extract between 30-60% (ImmunoCAP) and reduced peanut extract-induced skin wheal sizes by 1.6-7.4 millimeters in peanut allergic patients. We created a molecular map of the IgE antibody response to the most important peanut allergen proteins to enable the design of new allergy immunotherapies and vaccines.

Bacillus thuringiensis Cry1A Insecticidal Toxins and Their Digests Do Not Stimulate Histamine Release from Cultured Rat Mast Cells.

Public acceptance of genetically modified crops engineered with Bacillus thuringiensis (Bt) insecticidal protein genes (BT-GMCs), which confer resistance to various lepidopteran insect pests, is generally lacking. As a major concern over BT-GMCs is the allergenicity of insecticidal proteins, alleviating safety concerns should help increase public acceptance. In this study, three lepidopteran-specific Bt toxins, Cry1Aa, Cy1Ab, and Cry1Ac, were treated with simulated digestive fluids under various conditions. Western blotting using antiserum raised against individual segments (α-helices of domain I and β-sheets of domains II and III) of Cry1Aa showed that digestion produces a variety of polypeptides. In particular, the transmembrane α4-α5 of domain I, which may retain the ability to form pores, was the most resistant to digestion. Intact Cry1A toxins and these digests were then applied to RBL-2H3 cultured rat mast cells to determine whether the toxins directly induce histamine release. However, fluorescence microscopy revealed no specific binding of Cry1A toxins to RBL-2H3 cultured rat mast cells. In addition, neither the OPA method nor HPLC analysis detected significant histamine release from mast cells treated with Cry1A toxins and these digests. Our results provide important data supporting the safety of Cry1A toxins and potentially BT-GMCs.

Extracellular Vesicles Are Prevalent and Effective Carriers of Environmental Allergens in Indoor Dust.

The global incidence of allergic diseases is rising and poses a substantial threat to human health. Allergenic proteins released by various allergenic species play a critical role in the pathogenesis of allergic diseases and have been widely detected in the environmental matrix. However, the release, presence and interaction of environmental allergens with human body remain to be elucidated. In this study, we reported the widespread of allergen-harboring extracellular vesicles (EVs) in indoor dust from 75 households across five provinces in China. Particle size and abundance of EVs were correlated with specific environmental factors. EVs showed long persistence and high resistance to environmental stress. Metagenomics and metaproteomics data revealed that most indoor allergenic species released allergens within the EVs into dust. A higher abundance of allergenic species and their derived EVs was observed in urban areas compared to rural areas. ELISA confirmed the allergenic activity of the EV-associated allergens. Allergens are common components and even markers of EVs, as evidenced by the data compilation of various allergenic species. The proportion of EV-associated allergens varied across species. EVs facilitated allergen entry into epithelial cells. Intranasally administered EVs can be rapidly transported to the lungs and gastrointestinal tract. EV-associated allergens exhibited higher allergenicity compared with non-EV allergens. Our findings elucidate a vesicle pathway through which environmental allergens are released, persist, and trigger allergic responses within EVs.

Rebalancing the seed proteome following deletion of vicilin-related genes in pea (Pisum sativum L.).

Null mutations for genes encoding a major seed storage protein in pea, vicilin, were sought through screening a fast-neutron mutant population. Deletion mutations at four or five vicilin loci, where all vicilin genes within each locus were deleted, were combined to address the question of how removal or reduction of a major storage protein and potential allergen might impact the final concentration of protein per unit mature seed weight, seed yield and viability. While the concentration of seed protein was not reduced in mature seeds of mutant lines, indicative of a re-balancing of the proteome, notable differences were apparent in the metabolite, proteomic and amino acid profiles of the seeds, as well as in some functional properties. Major effects of the deletions on the proteome were documented. The genomic regions which were deleted were defined by whole genome sequencing of the parental line, JI2822 and its quintuple vicilin null derivative, providing a comprehensive description of each vicilin locus and its genic arrangement. An annotated reference genome has been generated for JI2822, which will serve as a very valuable resource for the research community and support further study of the associated deletion mutant population.

Assessment of transglutaminase catalyzed cross-linking on the potential allergenicity and conformation of heterologous protein polymers.

Transglutaminase (TGase)-mediated cross-linking has gained significant attention due to its potential to reduce the allergenicity of food proteins. This study investigates the effects of TGase cross-linking on allergenicity and conformational modifications in a dual-protein system comprising soy protein isolate (SPI) and β-lactoglobulin (β-LG). The results showed that TGase cross-linking effectively decreased the allergenic potential of both SPI and β-LG, with a more pronounced reduction observed in the allergenicity of soy protein in the dual-protein system. SDS-PAGE analysis revealed that the 7S and 11S subunits of soy protein were more easily cross-linked than β-LG. Secondary structure analysis indicated that TGase treatment disrupted β-sheet structures, increased the content of random coils, and enhanced protein flexibility. Ultraviolet absorption and intrinsic fluorescence analyses confirmed these structural alterations, with TGase treatment exposing additional aromatic amino acids. A reduction in free sulfhydryl groups and altered intermolecular forces further corroborated the occurrence of cross-linking. These findings suggest that TGase-mediated cross-linking effectively reduced the allergenicity of SPI and β-LG by modifying their conformations, offering potential strategies for the development of hypoallergenic dual-protein food products. PRACTICAL APPLICATION: This study has practical applications in the food industry to develop hypoallergenic food products, particularly those that combine soy and dairy proteins. By using TGase to cross-link these proteins, the allergenicity can be reduced, resulting in products that are safer for consumers with food allergies.

Diagnosis of soybean and peanut allergen components: identification of major components and clinical management strategies

Soybeans and peanuts belong to the leguminous family and are common causes of food anaphylaxis. Symptoms range from oral allergy syndrome to severe breathing difficulties, anaphylactic shock, and even death. But the allergens causing allergies are different, and the severity of symptoms are different. Precise diagnosis at the molecular level or component-resolved diagnosis (CRD) is necessary. CRD is a technology that accurately identifies allergen proteins to help physicians personalize clinical treatment and management strategies for allergy patients. In this article, according to the European Academy of Allergy and Clinical Immunology (EAACI) issued the "Allergen Component Diagnostic Guidance Recommendation 2.0 (MAUG 2.0)", it introduces the soy and peanut allergenic components, clinical diagnosis, treatment and management, aimed at improving the accurate diagnosis and personalized treatment of soy and peanut allergy.

Mitigating Food Protein Allergenicity with Biopolymers, Bioactive Compounds, and Enzymes

This review explores strategies for mitigating food allergies by treating foods with biopolymers, bioactive compounds, and food-grade enzymes. Biopolymers like chitosan, alginate, and pectin show potential in reducing the allergenic properties of food. Polyphenols such as quercetin, resveratrol, curcumin, and epigallocatechin gallate demonstrate promise as anti-inflammatory molecules that can lessen the symptoms and severity of allergic reactions. Enzymes, including proteases such as pepsin, papain, and bromelain, and transferases like transglutaminase, offer the potential to reduce the allergenic potency of proteins by various mechanisms, though more research is needed for the optimization and assessment of the safety and palatability of treated foods. Overall, this review offers insights into potential strategies to alleviate allergic reactions by reducing the allergenic properties of food proteins.

Review of Methods for the Detection of Allergens in Novel Food Alternative Proteins

Novel food, defined in the retained food regulations as ‘food that has not been consumed to a significant degree by humans in the EU before 15 May 1997 (EU Regulation 2015/2283),’ is expected to be used to an increasing extent and in a range of food products within the coming decade aiming to feed the growing global population in a more sustainable manner and comprising nutritious forms of protein. The safety aspects of novel foods must be thoroughly assessed before they can reach the market, and this includes assessment of allergenicity risks. FSA have funded this project to review current knowledge of the allergenicity of insect protein and precision fermentation (PF) protein and identify future research needs in this field. This report comprises an unbiased critical literature review (Section 1) coupled with consultations with experts and stakeholders in the field (Section 2) and focussing specifically on insect protein as well as milk and egg protein produced by precision fermentation. Section 3 comprises testing data to determine whether current allergen testing ELISA kits can be used to detect allergens in novel food. The literature review addresses allergenicity considerations of these novel proteins. The expert consultation (allergen testing, innovative methods, protein biochemistry) aimed to gain information on the potential allergenicity of PF and insect proteins, cross-reactivity (insect/shellfish allergens) and to identify knowledge gaps and challenges to recommend future strategies.

Identifying putative allergens from Cenostigma pluviosum pollen using proteomic bioinformatics.

Routinely, signs and symptoms of pollinosis worsen during the flowering period of the Cenostigma pluviosum var. peltophoroides (Sibipiruna) trees. This study aimed to determine whether Sibipiruna pollen contains allergenic proteins. The pollen from mature Sibipiruna anthers was morphologically characterized using optical, scanning electron and emission microscopy. Additionally, biochemical characterization was conducted through Shotgun Label-Free Proteomic Analysis. Three hundred and forty-nine proteins were identified using the UNIPROT database and thirty-six were confirmed as homologous allergenic proteins in the Allergome database. At the first time, Sibipiruna pollen allergenic proteins have been described and this study provides a deeper understanding of the allergenic proteins present in this pollen. These preliminary findings can be useful to assist in the development of targeted extracts for the study of sensitization and potential immunotherapy in the future.

Insight into the conformational changes and allergenic reactivity of coarse wheat bran induced by ozone treatment.

Whole grain food has gradually come into view because of its high nutritional value, but the incidence of wheat allergies has been increasing year by year. Ozone is a new non-thermal desensitization technology in food processing, and its effect on coarse wheat bran protein has received little study. We investigated the effects of different ozone airflow rates and treatment times on wheat bran allergenic property and protein structure. The ozone treatment time of 30 min at an airflow rate of 5 L min-1 reduced the coarse wheat bran allergenic property by 61.14%; subunits of 33, 46, 48 and 68 kDa were significantly less allergenic via western blotting; and the lowest releasing rate of β-hexosaminidase was 25.32% in the cell model of rat basophilic leukaemia 2H3. According to secondary structure and chemical interaction determination, the protein molecules were reorganized and aggregated by disulfide bonds and hydrophobic contacts following ozone exposure. Ozone treatment has a beneficial potential in reducing the allergenic property of coarse wheat bran and improving the safety of whole wheat products. © 2024 Society of Chemical Industry.

Impact of Heat and Pressure Processing Treatments on the Digestibility of Peanut, Hazelnut, Pistachio and Cashew Allergens.

Food processing can alter protein biochemical properties, impacting immunoreactivity and allergenicity. A key feature of food allergens is their resistance to enzymatic digestion, particularly by pepsin and trypsin. This study compares the digestomes of raw and heat- and/or pressure-treated peanuts, hazelnuts, pistachios and cashews using the INFOGEST harmonized digestion protocol and analyzing their IgE-binding capacity through in vitro methods. Protein patterns from controls and digestomes were resolved by SDS-PAGE and tested with sera from allergic patients, confirmed by competitive ELISA for hazelnuts and peanuts. The results indicate that processing methods differently affect the gastrointestinal (GI) digestion of these allergens. Simulated GI digestion caused a significant destruction of protein structures, reducing but not eliminating IgE reactivity for all four nuts. Boiling for 60 min did not change the SDS-PAGE profiles, but it did stimulate enzymatic activity, decreasing IgE binding capacity. In contrast, applying heat and pressure led to a nearly complete inhibition of allergenic potential during simulated digestion. These findings suggest that employing intense food processing techniques and investigating the gastrointestinal effects of highly allergenic nuts could be crucial steps toward developing new hypoallergenic formulations.

Pathogenesis-related protein, thaumatin-like protein 1b is a transdermal sensitizing allergen of mango fruits in mouse models

ABSTRACT Food allergens can enter the skin and increase the susceptibility to food allergies by producing immunoglobulin (Ig) E. Mango is a popular fruit worldwide and known to be allergenic. Dermal sensitization may occur around the oral cavity during ingestion of mangoes. However, to date, no allergenic proteins in mangoes have been shown to cause dermal sensitization in humans or animal models. Therefore, we aimed to identify mango proteins that sensitize mice percutaneously. The occiput of BALB/c mice was shaved, and a crude mango extract containing sodium dodecyl sulfate was applied to the skin. Mango protein-specific IgE and IgG1 were produced in response to this treatment. Proteins in mangoes that bind to mouse IgG1 and IgE were purified and identified as thaumatin-like protein 1b (TLP1b). This is the first study to identify TLP1b, a pan-allergen belonging to a family of pathogenesis-related proteins, as a mango allergen causing transdermal sensitization in mice.