What makes a food protein an allergen?

Food allergens are almost always proteins, but not all food proteins are allergens. This one statement sums up the purpose of this article, defining the difference between an innocuous food protein and a food allergen. The simplest answer is that a food allergen has the ability to first elicit an IgE response, and then, on subsequent exposures, to elicit a clinical response to the same or similar protein. However, this simplistic answer avoids the more complex issues of defining the biochemical characteristics that allow a food protein to survive the extremes of food processing, escape the digestive enzymes of the human gastrointestinal tract, and interact with the immune system. More than 700 allergen sequences have been identified from food and nonfood sources. However, despite increasing knowledge of the structure and amino acid sequences of the identified allergens, only a few biochemical characteristics can be associated with food allergens. Food allergen characteristics, including abundance of the protein in the food; multiple, linear IgE binding epitopes; resistance of the protein to digestion and processing; and allergen structure are discussed, and the possible reasons they predispose some food proteins to become allergens are suggested.     

Meeting the nutritional needs of the low-birth-weight infant

Approximately 5% of young children and 3-4% of adults exhibit adverse immune responses to foods in westernized countries, with a tendency to increase. The pathophysiology of food allergy (FA) relies on immune reactions triggered by epitopes, i.e. small amino-acid sequences able to bind to antibodies or cells. Some food allergens share specific physicochemical characteristics that allow them to resist digestion, thus enhancing allergenicity. These allergens encounter specialized dendritic cell populations in the gut, which leads to T-cell priming. In case of IgE-mediated allergy, this process triggers the production of allergen-specific IgE by B cells. Tissue-resident reactive cells, including mast cells, then bind IgE, and allergic reactions are elicited when these cells, with adjacent IgE molecules bound to their surface, are re-exposed to allergen. Allergic reactions occurring in the absence of detectable IgE are labeled non-IgE mediated. The abrogation of oral tolerance which leads to FA is likely favored by genetic disposition and environmental factors (e.g. increased hygiene or enhanced allergenicity of some foods). For an accurate diagnosis, complete medical history, laboratory tests and, in most cases, an oral food challenge are needed. Noticeably, the detection of food-specific IgE (sensitization) does not necessarily indicate clinical allergy. Novel diagnostic methods currently under study focus on the immune responses to specific food proteins or epitopes of specific proteins. Food-induced allergic reactions represent a large array of symptoms involving the skin and gastrointestinal and respiratory systems. They can be attributed to IgE-mediated and non-IgE-mediated (cellular) mechanisms and thus differ in their nature, severity and outcome. Outcome also differs according to allergens.     

Can any protein become an allergen?

Allergens of plant and animal foods and pollen belong to a highly restricted number of protein families. The AllFam Database (http://www.meduniwien.ac.at/allergens/allfam/) provides regularly updated lists of protein families that contain allergens. At present, 2% of the 9318 protein families defined by the Pfam Database (http://pfam.sanger.ac.uk/) contain allergens. Related protein families can be grouped into superfamilies placing allergenic proteins in an evolutionary context. With the exception of the prolamin superfamily, allergenic plant proteins are found in few member families of their respective superfamilies. This might indicate that allergenicity emerged rather infrequently in a very limited number of protein families. Moreover, most members of a given protein family seem to be non-allergenic. In contrast to plant allergens, the allergenicity of animal food allergens seems to be dependent on the degree of identity to a human homologue. The closer a potential animal allergen is to a human protein, the less likely it is to act as allergen.     

Selective hypersensitivity to boiled razor shell.

Many types of seafood require cooking before ingestion and it has been demonstrated that this cooking process may affect the antigenicity and allergenicity of the food. We describe a case of anaphylaxis caused by selective sensitization to razor shell, a mollusc. In vivo and in vitro studies confirmed sensitization to boiled razor shell. Analysis of the nature of the allergen yielded results that were consistent with the findings of other authors and suggested that allergens involved in seafood allergy are commonly high molecular weight proteins that, in most cases, are heat stable.     

Factors influencing the allergenicity and adjuvanticity of allergens

IgE-mediated allergic disorders affect up to 25% of the population in industrialized countries and result in a Th2-polarized immune response to innocuous environmental proteins, so-called allergens. Among a large number of proteins to which humans are exposed to, only a minute fraction are allergens. This observation suggests that allergens share special features of allergenicity (i.e., the capacity to induce the production of specific IgE antibodies in susceptible individuals). However, the question 'what makes a protein allergenic' still remains unanswered although some biochemical characteristics of allergens and their capacity to interact with the innate immune system could be associated with their allergenic potential. Allergen-specific immunotherapy aims at an alteration of the disease-eliciting immune response by repeated administration of allergens. Recently, approaches emerged to endow allergens with adjuvanticity, in particular aiming at an increase of their immunomodulatory capacity. This article summarizes factors of allergenicity and introduces recent concepts of adjuvanticity to improve allergen-specific immunotherapy.     

Vaccines and Immunomodulatory Therapies for Food Allergy

The apparent increase in food allergy prevalence has led to a surge in the amount of clinical and basic science research dedicated to the field. At the current time, allergen avoidance remains the cornerstone of treatment; however, recent clinical trials investigating various forms of immunotherapy have opened doors to the possible future application of an active treatment strategy in everyday practice. In addition, improvements in molecular biology have allowed researchers to purify, clone, and modify allergens, thus laying the groundwork for research on vaccines using modified proteins of decreased allergenicity. Finally, various allergen-nonspecific immunomodulatory therapies are also being investigated as a means to alter the immune response to food allergens. With these emerging therapeutic strategies, it is hoped that practitioners will have options in caring for their food-allergic patients in the near future.     

Pathophysiology of food-induced anaphylaxis

Food-induced anaphylaxis is a steadily increasing problem in westernized countries and now represents the leading cause of anaphylaxis in the outpatient setting, particularly in children. Much of our knowledge of the pathophysiology of food-induced anaphylaxis comes from animal studies. Food anaphylaxis in humans is thought to be entirely IgE mediated. Several features appear to be unique to these reactions; factors such as exercise can lower the “threshold” for anaphylaxis in certain susceptible individuals. Different methods of thermal processing can modify the allergenicity of food proteins. Alteration of stomach pH can allow for incomplete digestion of food proteins, leading to increased absorption of intact food allergens. Low serum platelet-activating factor acetylhydrolase may predispose to fatal food-induced anaphylaxis. With a greater understanding of the pathophysiology of food-induced anaphylaxis, novel approaches not only to identify those at risk, but to treat and ultimately prevent food-induced anaphylaxis, are on the horizon.     

Les plantes transgéniques (OGM végétaux) : connaissances et inconnues sur les risques d'allergénicité…

Much attention is now being focused on foods from genetically modified plants because of the risk of allergenicity. No such risk has been reported for the first generation of GM plants made resistant to herbicides and insect larvae. Current experiments with hypoallergenic GM plants are reported and discussed in the present paper. The second generation of GM plants will improve the nutritional aspects of natural foods. Transgenic proteins could reach from 4 up to 8% of the total protein content in these foods. Any potential difference in allergenicity between second generation GM plants and the natural varieties must be examined with respect to the risk for food allergy caused by food products made from these plants and the risk for respiratory allergies in the people living near the crops caused by airborne pollen originating from the plants. WHO–FAO directives as well as the Codex Alimentarius proposals and the European Food Safety Authority (EFSA) guidelines recommend that transgenic proteins be screened for homology (by in silico study) and cross-reactivity with known allergens, as well as being examined carefully for modifications of host-plant proteomes. In vivo animal studies are also to be carried out to assess any potential immunogenicity. Lacking adequate safety data, the absence of potential allergenicity of transgenic plants cannot be ruled out. This is why data that do not meet the recommended safety criteria required for commercialization of GM plants do not allow us to rule out absolutely the risk that may be associated with products that are going to be commercialized. Therefore, it is essential that commercialized GM plants be monitored. We propose the establishment of public reference serum banks based on up-to-date WHO–FAO recommendations concerning the selection of sera according to precise criteria. We also propose establishing a system of allergovigilance linking national and European health and food safety agencies and a network of university hospital-based clinical and laboratory reference centres, together with a network of clinical allergists, responsible for the creation of the serum banks. Allergists working through these networks would be able to identify new sensitizations to transgenic foods in the population, just as they now identify new types of food allergies, which, in this case, would be GM foods. Such a project is now being established in France.     

Animal models for assessment of GMO allergenicity: advantages and limitations

Incidence of IgE-mediated allergic reactions to foods is increasing as well as the severity of associated symptoms and numerous foods are now incriminated, probably in relation with modifications of dietary habits and increased exposure to new or modified food ingredients. Therefore, the introduction on the market of food composed of or derived from genetically modified organisms (GMOs) raised the question of their potential allergenicity. Particularly with regards to the allergenicity of a newly expressed protein, it is necessary to obtain, from several steps in the risk assessment process, a cumulative body of evidence which minimises any uncertainty. This may include the use of animal model despite no fully reliable validated model is available yet. Such animal models should allow to address 3 major issues: Is the novel protein a sensitizer, i.e. does it possess intrinsic properties that allow to sensitize a predisposed individual? Is the protein an elicitor i.e. is it able to elicit an allergic reaction in a sensitised individual? And is the protein an adjuvant, i.e. can it facilitate or enhance the sensitisation to an other protein? Animal models under investigation currently include mice, rats and guinea pigs but models such as dogs and swine also appeared a few years ago. The aim is to mimic the mechanism and characteristics of the sensitisation phase and/or the elicitation phase of the allergic reaction as it occurs in atopic humans. They are necessary because sensitisation studies can obviously not be done in human and because in vitro tests cannot reproduce the complexity of the immune system. We propose a mouse model which mimics both phases of the allergic reaction. It has permitted to evidence that biochemical and clinical manifestations occuring during the active phases of the allergic reaction differ according to the structure of the allergen used for the challenge. This may allow to compare the allergenic potential of a genetically modified protein with that of the conventional one and to identify possible unintended effects. However, pathogenesis of food allergy in human is very complex and multifactorial, including individual differences in susceptibility, environmental factors, conditions of exposure, ... No animal model can take into account all these factors and allow a reliable prediction of the prevalence and severity of allergic reactions which would result from the exposure to a (novel) protein. Nevertheless, point by point analysis using the different models available may provide useful informations on the potential allergenicity of a novel protein.     

Studies on BN rats model to determine the potential allergenicity of proteins from genetically modified foods

AIM: To develop a Brown Norway (BN) rat model to determine the potential allergenicity of novel proteins in genetically modified food. METHODS: The allergenicity of different proteins were compared, including ovalbumin (OVA), a potent respiratory and food allergen, bovine serum albumin (BSA), a protein that is considered to have a lesser allergenic potential, and potato acid phosphatase (PAP), a non-allergenic protein when administered to BN rats via different routes of exposure (intraperitoneally or by gavage). IgG and IgE antibody responses were determined by ELISA and PCA, respectively. An immunoassay kit was used to determine the plasma histamine level. In addition, possible systemic effect of allergens was investigated by monitoring blood pressure. RESULTS: OVA provoked very vigorous protein-specific IgG and IgE responses, low grade protein-specific IgG and IgE responses were elicited by BSA, while by neither route did PAP elicit anything. In either routes of exposure, plasma histamine level in BN rats sensitized with OVA was higher than that of BSA or PAP. In addition, an oral challenge with BSA and PAP did not induce any effect on blood pressure, while a temporary drop in systolic blood pressure in few animals of each routes of exposure was found by an oral challenge with OVA. CONCLUSION: BN rat model might be a useful and predictive animal model to study the potential allergenicity of novel food proteins.     

Airborne allergy to sunflower seed.

BACKGROUND: There is increasing evidence that bird fanciers may develop airborne allergies to unusual allergens. OBJECTIVE: To detect the allergen source in a bird fancier with a history of asthma associated with bird cage cleaning activities and with contact with a Brazil parrot. METHODS: SPT with a large series of both airborne and food allergens were carried out. IgE reactivity to allergens causing wheal and flare reactions was confirmed by in-vitro investigations including ELISA/ELISA inhibition and immunoblot analysis. RESULTS: Strong skin reactivity to sunflower seed was observed. Immunoblot analysis showed IgE reactivity to low m.w. proteins, most probably 2S albumin, and ELISA inhibition studies showed the absence of cross-reactivity to mustard. CONCLUSION: Sunflower seed dust may sensitize patients via the respiratory tract. Differently from previously reported cases of sunflower seed allergy, no cross-reactivity to 2S albumin from botanically unrelated seeds was found.     

Allergies associated with both food and pollen

Recent progress in understanding structural relationships between allergens has allowed their classification into molecular families. Proteins belonging to a molecular family often show some degree of IgE cross-reactivity. These cross-reactions can lead to a clinical association like birch-apple syndrome whose basis is a sensitization to a PR-10 protein (birch pollen Bet v 1) and then oral symptoms in contact to apple Mal d 1, another PR-10 family member. Food allergens implicated into pollen-food allergy syndromes differ from those linked to crustacea or milk cross-allergies: they seem unable to sensitize the patient through oral route. As a result, they most often induce weaker clinical reactions than complete allergens like those present in shrimp or cow milk. Numerous molecular families have been isolated from pollens. PR-10 and profilins have a well established role in inducing clinical reactions to food like fruits and vegetables. Some molecular families need more studies to delineate their true impact on pollen-driven food reactions: polygalacturonases, pectate lyases, isoflavone reductases, thaumatin-like, cyclophilins.... Others are found in pollen but not in eaten products: 2-EF-hand calcium binding proteins, beta expansins,... Lipid transfer proteins (LTP) are widespread plant food allergens (e.g. in peach): these proteins seem able to directly sensitize the patient through oral route. But recent data have suggested a possible additional effect of some LTP present in pollens (mugwort, olive, pellitory).     

Les allergènes recombinants : leur apport à l’allergologie en 2006

Advances in molecular biology techniques have led to the production of recombinant allergens, about thirty of them now being available for measurements (DIAGNOSIS?) in vitro. These recombinant allergens correspond to a precise molecular variant of a natural allergen, and their biological activity has to be evaluated in comparison with the corresponding natural allergen. The advantages of recombinant allergens are essentially the creation of allergenic preparations having constant pharmaceutic properties, which allows determination of specific IgE directed against different molecular components of an allergenic source, for example, pollen, mites, etc. The main consequences of these biological advances are the following: evaluation of sensitivities to allergen molecules in different populations (molecular epidemiology), improvement of extracts used for diagnosis by selection of the most pertinent allergenic sources and in quantifying their major allergen content, definition of the spectrum of recognition of specific IgE vis-à-vis different molecular components (spectrotype), quantitative evaluation of IgE responses, establishing the molecular basis of cross-reactions between different inhaled allergens, between different food allergens, and between inhaled allergens and food allergens. As regards allergy practice, this new diagnostic tool can lead to better interpretation of polysensitivities, observed by skin tests and in vitro tests. Some examples of particular clinical cases associated with specific sensitivities vis-à-vis certain recombinant allergens will be presented.     

Allergie respiratoire aux protéines de feuilles, implication d’un nouvel allergène

Symptoms of subjects presenting with rhinitis, conjunctivitis or contact urticaria when exposed or in contact with grass are usually attributed to allergy to grass pollen or to certain airborne molds. The four cases described in the present report presented with allergic symptoms when mowing their lawn. The suspicion of allergy to grass leaves was confirmed by skin prick tests with native leaves. An extract of rye grass leaves was made and its allergens were analyzed by SDS-PAGE and immunoblotting. Three of the four patients were found to have IgE specific for a single 56 kDa molecule. It was shown to be a major leaf protein and identified as a subunit of ribulose 1,5 diphosphate carboxylase/oxydase, a major plant kingdom enzyme involved in photosynthesis. This protein is widely present in leaves and is, in addition, used as a non-allergenic model in investigation of the allergenicity of food proteins. In fact, it is degraded instantaneously by digestive enzymes, in contrast to the known principal food allergens. In conclusion, respiratory allergy to grass leaf proteins was demonstrated in this study of four patients, who were or were not allergic or sensitized to grass pollen.     

Food allergy: effect of proteins-lipids and proteins-polysaccharides interactions

The most widely used ingredients in food formulation are proteins, lipids and polysaccharides. Proteins-lipids and proteins-polysaccharides interactions play a key role in the structure, stability, sensorial and nutritional properties of formulated foods. The objective of the present study is to highlight the importance of proteins-lipids and proteins-polysaccharides interactions, on the immuno-reactivity of allergenic proteins. Two models have been studied, on the one hand refined and not refined oils (soya and sunflower) and soya lecithin, on the other hand mixtures based on peanut proteins and polysaccharides (arabic gum, pectin, xylan). STUDY OF OILS: We have extracted proteins, using a PBS buffer, from refined and not refined oils from soya, sunflower and from soya lecithin, determined protein concentrations and identified allergenic proteins using SDS-PAGE electrophoresis and immuno-blotting. Phospholipids are determined by atomic absorption spectrometry. The protein determination and SDS-PAGE show the presence of a higher amount of proteins in not refined oils and lecithin as compared to refined oils. An important amount of proteins associated to phospholipids are eliminated by degumming on the form of lecithin. On the other hand, residual proteins from refined oils are accompanied by phospholipids. Immuno-blots reveal the presence of a 56 kDa allergen in oils issued from soya seeds and soya lecithin, and the presence of a 67 kDa allergen in oils issued from sunflower seeds. We conclude that the presence or elimination of proteins, especially allergens from oils is linked to amphiphilic association to phospholipids. STUDY OF PEANUT PROTEINS-POLYSACCHARIDES MIXTURES: We have digested in vitro proteins in a dialysis bag using a multi-enzymatic method and characterized proteins and peptides using SDS-PAGE electrophoresis and immuno-blotting. Our results confirm that peanut proteins alone are digested by proteases and that a number of large peptides still have epitopes recognized by anti-peanut proteins antibodies. Our results also show that the presence of polysaccharides changes the peptidic profile after digestion and that, depending on the polysaccharide type, smaller or larger peptides can be obtained in the dialysis bag. Smaller peptides are obtained using pectin whereas larger peptides are obtained using arabic gum and xylan. In the latter case, an increasing amount of peptides reacts to antibodies. Our first observations clearly show the need to better understand modifications of proteins allergenicity induced by the presence of other ingredients such as polysaccharides and lipids, in relation to technological treatments.     

Animal models of food allergy: Opportunities and barriers

The potential for animal models to mimic the human disease process makes them an attractive tool for determining disease mechanisms, predicting disease triggers, and testing treatment regimens. With this in mind, animal models of food allergy have been receiving increasing attention as research tools to answer some of the difficult questions regarding food-allergy disease. Most of the food-allergy animal models developed to date have been designed to test reagents for immunotherapeutic treatment of allergic disease and to predict the potential human allergenicity of proteins. Current animal models under development are rodent, swine, and dog. The variables affecting development of such models include allergen concentration, allergen matrix or food source, allergen route of exposure, duration, animal age, adjuvant use, and dose range of allergens. Each model presents opportunities for and barriers to a fuller understanding of the allergic response. The conditions inherent to each model and the intended purpose of the study should therefore be considered prior to its use.     

Detection of four distinct groups of hen egg allergens binding IgE in the sera of children with egg allergy

BackgroundThere appears to be a lack of agreement in the literature on the allergenicity of hen egg proteins. This may be partly due to the use of impure proteins in some cases. Egg yolk proteins have also been largely ignored in such studies. We therefore set out to determine, using especially purified proteins, their relative allergenicity, and to observe whether there were any relationships between their potency and the sensitivity of patients to them.Methods and resultsThe sera of 40 patients with clinically observed hen egg hypersensitivity were tested for specific IgE binding to purified egg white and egg yolk proteins using the radioallergosorbent test (RAST). Statistical treament by correspondence analysis of the percent radioactive uptakes in the RAST to the 8 proteins demonstrated that there were four distinct groups of patients reacting in a similar way to four discrete sets of proteins.ConclusionsThe first three sets of allergens consisted of egg white proteins as follows: firstly, lysozyme and ovalbumin; secondly, ovomucoid; and thirdly, ovomucin. The fourth set contained the egg white protein ovotransferrin and the egg yolk proteins apovitellenins I and VI and phosvitin. The existence of patient groups may explain why various workers have reported different allergens to be important in egg hypersensitivity. A sufficiently large number of patients must be examined so as to give a representative distribution across each group, otherwise the results may be biased towards one allergen.     

Overview of Component Resolved Diagnostics

Component-resolved diagnostics (CRD) utilize purified native or recombinant allergens to detect IgE sensitivity to individual allergen molecules and have become of growing importance in clinical investigation of IgE-mediated allergies. This overview updates current developments of CRD, including multiarray test systems. Cross-reactions between allergens of known allergen families (i.e. to Bet v 1 homologues) are emphasised. In pollinosis as well as in allergy to hymenoptera venoms or to food, CRD allows to some extent discrimination between clinically significant and irrelevant sIgE results and the establishing of sensitisation patterns with particular prognostic outcomes (i.e. sensitisations to storage proteins which correlate with clinically severe reactions in peanut allergy). Further promising improvements in diagnostics are expected from additional, not yet commercially available, recombinant allergen diagnostics identifying particular molecules of risk. Overall, CRD may decrease the need for provocation testing and may also improve the specificity of allergen-specific immunotherapy.     

Evaluation of Molecular Basis of Cross Reactivity between Rye and Bermuda Grass Pollen Allergens

BackgroundAllergenic cross reactivity between the members of the Pooids (Lolium perenne, Phleum pratense, and Poa pratensis) and Chloridoids (Cynodon dactylon and Paspalum notatum) is well established. Studies using crude extracts in the past have demonstrated limited cross reactivity between the Pooids and the Chloridoids suggesting separate diagnosis and therapy. However, little is known regarding the molecular basis for the limited cross reactivity observed between the 2 groups of grasses. The present study was undertaken to gain insights into the molecular basis of cross allergenicity between the major allergens from rye and Bermuda grass pollens.MethodsImmunoblot inhibition tests were carried out to determine the specificity of the proteins involved in cross reactivity. Crude pollen extract and bacterially expressed and purified recombinant Lol p 1and Lol p 5 from rye grass were subjected to cross inhibition experiments with crude and purified recombinant Cyn d 1 from Bermuda grass using sera from patients allergic to rye grass pollen.ResultsThe immunoblot inhibition studies revealed a high degree of cross inhibition between the group 1 allergens. In contrast, a complete lack of inhibition was observed between Bermuda grass group 1 allergen rCyn d 1, and rye grass group 5 allergen rLol p 5. Crude rye grass extract strongly inhibited IgE reactivity to Bermuda grass, whereas crude Bermuda grass pollen extract showed a weaker inhibition.ConclusionsOur data suggests that a possible explanation for the limited cross reactivity between the Pooids and Chloridoids may, in part, be due to the absence of group 5 allergen from Chloridoid grasses. This approach of using purified proteins may be applied to better characterize the cross allergenicity patterns between different grass pollen allergens.     

The Molecular Basis of Peanut Allergy

Peanut allergens can trigger a potent and sometimes dangerous immune response in an increasing number of people. The molecular structures of these allergens form the basis for understanding this response. This review describes the currently known peanut allergen structures and discusses how modifications both enzymatic and non-enzymatic affect digestion, innate immune recognition, and IgE interactions. The allergen structures help explain cross-reactivity among allergens from different sources, which is useful in improving patient diagnostics. Surprisingly, it was recently noted that similar short peptide sequences among unrelated peanut allergens could also be a source of cross-reactivity. The molecular features of peanut allergens continue to inform predictions and provide new research directions in the study of allergic disease.