**3. Mechanism of action of allergenic proteins**

Out of the caseins that have been shown to elicit an IgE response in humans, the rule of thumb is the closer the sequence to the human equivalent is, the less likely an IgE response will occur. Conversely, sensitization to BSA is the main predictive marker for the cross-reactivity to cow's milk that is present in 73–93% of patients with beef allergies, despite of the fact that

Cow's milk and egg allergies are some of the most common food allergies found in young children. It is estimated that ~3.8 and 2% of children younger than 5 years old have cow's milk and egg allergies, respectively. These allergens are commonly found in a variety of foods due to their technological and nutritional importance [6]. Thus, lactoglobulin is the major allergen in cow´s milk. Milk contains more than 20 protein fractions. In the curd, four caseins account for ~80% of milk proteins. The remaining 20% of the proteins, are globular proteins (e.g., lactalbumin, lactoglobulin, and bovine serum albumin), and are found in whey and egg

Parvalbumin represents the major clinical cross-reactive fish allergen. It contains heat-resistant linear epitopes that are s sensitizing by the interaction of metal-binding domains. In addition, other fish allergens are found in collagen and gelatin isolated from skin and muscle tissues. On the other hand, in shellfish, crustaceans and mollusks, tropomyosin is the major allergen

About 65% of plant food allergens belong to the group of prolamins, termites and the group of proteins related to pathogenesis (PR-10) [25]. The prolamin family include storage proteins, lipid transfer (LTPs), alpha-amylase/protein inhibitors and albumins from most cereal seeds. Their 3D structure consists of a compact structure with four alpha-helices stabilized by disulfide bridges and a central cavity used for lipid binding. However, similarity in their third-party structures does not indicate a similarity in their amino acid sequence between proteins in the same group. Further, prolamins from wheat are known to cause baker's asthma and celiac disease in humans. Alpha-amylase/trypsin inhibitors from various cereals, such as wheat, barley and rice, have also been involved in allergies. Some of the clinically reported allergenic plant proteins belongs to the family of prolamins such as Ara h 2, Ara h 6, Sin a 1, Ber e 1, Ses i 2 and Jug v 1. Non-specific LTPs are known to be the main food allergens in the fruits of the *Rosaceae* family. The presence of specific IgE in LTP is considered a significant risk factor for allergy and may serve as a diagnostic marker [25]. The most relevant termite-type allergens are the 7S and 11S globulins. 7S globulins include Arah1, Jugr2 and Sesi3, whereas the 11S globulins include the Arah3, soy glycinins, Bere2, Cora9 and Fage1 [32]. The third structure of the proteins belonging to the termite group consists of a series of antiparallel β leaves associated with an α-helix forming a cavity [32]. This structure is also found in several lipocalin-like proteins involved in the transport of hydrophobic ligands, including milk β-lactoglobulin [32]. Further, plant prophylines share about 70% of the amino

All gliadin and glutenin protein fractions have been described as wheat grain allergens. The main properties of these proteins is to form a continuous viscoelastic network when flour is mixed with water to form a dough to be used in products such as bread, pastries, pasta,

proteins from both components (i.e., yolk and egg white) causing sensitization [30].

BSA shares 76% identity to its human homolog [27, 29].

that triggers allergic reactions [31].

48 Allergen

**2.2. Allergenic vegetable proteins**

acid sequence homology [32–34].

The prevalence of food allergy has been steadily increasing around the world. The relevant risk factors for food allergies are: (i) an increased use of antacids which is translated in a reduced digestion of allergens; (ii) a reduced consumption of omega-3 polyunsaturated fatty acids in the diet; (iii) a reduced consumption of antioxidants; (iv) genetics; (v) male gender; (vi) race, since it is increased among asian and black children as compared to white children; (vii) an increased hygiene; (viii) a northern climate; (ix) obesity, since this is an inflammatory state; (x) timing and route of exposure to foods (increased risk for delaying allergens with possible environmental sensitization); and (xi) vitamin D insufficiency [1, 39]. Most food allergens belong to only a limited number of proteins, and around 65% of plant food allergens belong to just four protein families, such as the prolamin, cupin, Bet v 1-like, and profiling, whilst animal food allergens can be classified into three main families named as tropomyosins, EF-hand proteins and caseins [40]. In general, children food allergies to milk, egg, wheat, and soy typically resolve during childhood, whereas allergies to peanut, tree nuts, fish, and shellfish are persistent. The prognosis also varies with disorder; for instance, food allergy related to eosinophilic esophagitis appears to have a relatively poor chance of resolution. For instance, the resolution rates are slow for allergies that have been commonly outgrown, such as those to milk, egg, wheat, and soy [39].

The human body has a series of physiological barriers for protection against foreign antigens. In the digestive system, these barriers are composed of two groups: (i) non-immunological such as the gastric acid, pancreatic enzymes, intestinal enzymes, mucus, the membrane of the microvilli, the mucosal layer and intestinal peristalsis), and (ii) immunological, such as IgA, IgE, IgM, IgG, lymphocytes, macrophages, Peyer's patches, intestinal secretory IgA and secretory IgA in breast milk [30]. Usually, when the immune system recognizes food proteins as a foreign body, immunoregulatory mechanisms are established that lead to the acquisition of tolerance. Alterations in these regulatory mechanisms alter the induction of tolerance, resulting in food allergy [30]. An allergy reaction require a complex interaction between the protein and the immune system [26]. The National Institute of Allergy and Infectious Diseases of the United States, identified four categories of immune-mediated adverse food reactions such as IgE-mediated, non–IgE-mediated, mixed, and cell-mediated reactions. The most prevalent non-IgE-mediated reactions are eosinophilic esophagitis (EoE), the food-protein induced enterocolitis syndrome, proctocolitis, entheropathy and celiac disease [41]. The IgE-mediated reaction is by far, the most well established mechanism, where the antibodies bind to the high affinity receptors of mast cells and basophils, and to the low affinity receptors on macrophages, monocytes, lymphocytes and platelets. Thus, IgE are able to bind a specific receptor on the surface of mast cells and basophils, when two or more of these captive IgE molecules bind to their specific antigen, becoming cross-linked on the surface of the cell [26]. Once the allergens penetrate the mucosal barriers and bind to the IgE of mast cells and basophils, these cells release mediators that cause vasodilatation, smooth muscle contraction and mucosal secretion, giving rise to the typical symptoms of immediate hypersensitivity (**Annex 1**) [30].

In order to crosslinking to takes place, at least two antibody molecules must bind to the inducing allergen. An allergen must therefore contain at least two IgE binding sites, each one contains at least 15 amino acid residues. This implies a lower size limit for protein allergens of approximately 30 amino acid residues [26]. The IgE-mediated allergic immune response can be divided into three phases: (i) the sensitization phase in which B lymphocytes switch to the production of specific IgE, (ii) the effector phase consisting of an acute reaction and a facultative late-phase reaction; (iii) a chronic phase that may be the result of repetitive late phase reactions. The acute reaction causes activation of mast cells and basophils releasing histamine, leukotrienes, and other mediators known to be responsible for the wheal and flare reaction occurring in the skin and at various mucosal sites including the eye, nose, lung, and gastrointestinal tract [7]. The IgE-mediated reaction occur immediately or within 1–2 h of ingestion, whereas non-IgE–mediated reactions generally have a delayed onset beyond 2 h of ingestion [42].

A limited number of foods are responsible for the majority of reactions in IgE-mediated food allergy. For instance, allergy to cow milk, eggs, wheat, and soy are more common

**Annex 1.** IgE-mediated protein hypersensitivity.

in infants and young children, whereas seafood, peanuts, and tree nuts are the most common causes of food allergy in adults [40]. The IgE-mediated reaction in skin includes hives and angioedema, whereas the gastrointestinal manifestations include mouth and lip pruritus, abdominal pain, vomiting, and diarrhea. On the other hand, a variety of respiratory tract symptoms that generally involve IgE-mediated responses, includes rhinorrhea and wheezing, whereas isolated asthma or rhinitis are unusual [42]. Further, the IgE-mediated food allergy may cause the dietary protein–induced syndromes such as enteropathy and enterocolitis. These in turn, cause profuse vomiting, diarrhea, dehydration and lethargy. Other syndromes include proctocolitis, gastroesophageal reflux, infantile colic, constipation and the Heiner syndrome [42].
