Identification of an IgE Reactive Peptide in Hen Egg Riboflavin Binding Protein Subjected to Simulated Gastrointestinal Digestion

Martos, Gustavo; Pineda-Vadillo, Carlos; Miralles, Beatriz; Alonso-Lebrero, Elena; López‐Fandiño, Rosina; Molina, Elena; Belloque, Josefina

doi:10.1021/jf3001586

Cited by 9 publications

(11 citation statements)

References 23 publications

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“…6B). In the direct DD, and in agreement with their specificities, trypsin hydrolyzed LYS after Lys (1,13,33,96,97) and Arg (21,45,61,73,114); and chymotrypsin hydrolyzed LYS after Tyr (20,23,53), Trp (28,62,63,108,123), Phe (3,34,38), Leu (56, 75, 83, 84), and Met (12). There were also peptides corresponding to the cleavage of bonds with Asp (18), Asn (37, 74, 93), and Cys (76, 80) ( Fig.…”

Section: The Gastric and Gastroduodenal Digestion Products Of Lys Retsupporting

confidence: 55%

“…After SDS-PAGE separation on Tris-Tricine gels, these were soaked in transfer buffer (48 mM Tris, 39 mM glycine, 20% methanol, pH 9.2) for 20 min and subjected to semidry transfer in a Trans-Blot SD (Bio-Rad) for 20 min at 18 V. Then, the 0.22-m nitrocellulose membranes were developed according to [12] with pooled or individual sera (Supporting Information Table 1).…”

Section: Western Blottingmentioning

confidence: 99%

“…For intact molecular weight determination, proteins were electroblotted and directly analyzed after dissolution of the nitrocellulose membrane in MALDI matrix solution prepared as a saturated solution of ␣-CHCA in ACN/methanol 70/30% containing 1% TFA [12]. Samples were spotted on a Bruker Anchorchip target and spectra were acquired on linear mode.…”

Section: Maldi-tof/tof Analyses Of the Peptide Bandsmentioning

confidence: 99%

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Immunological behavior of in vitro digested egg‐white lysozyme

Jiménez‐Saiz

Benedé

Miralles

et al. 2013

Molecular Nutrition Food Res

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Scope Besides its antimicrobial properties, lysozyme (LYS) is one of the major allergens from hen egg. This paper addresses the identification of the peptides produced upon in vitro gastrointestinal digestion of LYS, together with their IgE‐binding and biological activity as a contribution to the understanding of what makes it a relevant allergen. Methods and results Simulated in vitro gastrointestinal digestion together with IgE binding, basophil degranulation, and peripheral blood mononuclear cells stimulation experiments were carried out. Identification of the fragments released was performed by HPLC‐MS/MS and the immunoreactive products were analyzed by MALDI‐TOF/TOF. Results showed that in vitro gastric and gastroduodenal digests of LYS maintained IgE binding, basophil activation capacity, and preserved T‐cell immunogenicity. These biological activities could be attributed to either the persistence of intact LYS, due to incomplete gastric degradation and subsequent duodenal precipitation, the formation of fragment f(24–129) by chymotrypsin action on the soluble intact protein, or the release, upon combined gastric and pancreatic digestion, of immunoreactive peptides linked by disulphide bonds containing the epitopes f(57–83) and f(108–122). Conclusion The pH of gastric hydrolysis greatly determined the extent of subsequent duodenal digestion of LYS and the disclosure of relevant epitopes that could increase its allergenic potential.

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Section: The Gastric and Gastroduodenal Digestion Products Of Lys Retsupporting

confidence: 55%

Section: Western Blottingmentioning

confidence: 99%

Section: Maldi-tof/tof Analyses Of the Peptide Bandsmentioning

confidence: 99%

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Immunological behavior of in vitro digested egg‐white lysozyme

Jiménez‐Saiz

Benedé

Miralles

et al. 2013

Molecular Nutrition Food Res

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“…After SDS-PAGE separation, the gels were soaked in transfer buffer (48 m M Tris, 39 m M glycine, 20% methanol, pH 9.2) for 20 min and subjected to semidry transfer in a Trans-Blot SD (Bio-Rad) for 20 min at 18 V. Then, 0.22-µm nitrocellulose membranes were developed according to Martos et al [12]. Sera from the above-described patient were used at different time points before and after EOIT, as well as individual serum samples from 5 children with proven allergy to egg proteins, diagnosed on the basis of clinical allergic symptoms of an acute reaction after egg ingestion, together with evidence of high LYS-specific IgE antibodies (from 0.59 to 266 kU/l) and a pool of 6 sera (on average 0.38 kU/l LYS-specific IgE).…”

Section: Methodsmentioning

confidence: 99%

Anaphylaxis Induced by a Drug Containing Lysozyme and Papain: Influence of Papain on the IgE Response

Benedé

Pérez-Rangel

Lozano‐Ojalvo

et al. 2014

Int Arch Allergy Immunol

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Background: This paper reports the case of an egg-allergic pediatric patient who, once desensitized to egg following a successful rush oral immunotherapy protocol, could also tolerate Lizipaina®, a drug containing lysozyme (LYS) and papain, which had previously caused him a severe allergic reaction. Because the LYS amount that elicited the anaphylactic reaction (5 mg) was much lower than that tolerated during a double-blind placebo-controlled food challenge (corresponding to approximately 60 mg of LYS), the possibility that the presence of papain could increase the allergenic potential of LYS was investigated. Methods: Lizipaina, LYS and LYS hydrolyzed with papain were analyzed by SDS-PAGE under reducing and nonreducing conditions, and Western blotting of sera from egg-allergic patients was performed in order to detect IgE-binding fragments. Finally, sequence identification of the IgE-reactive bands was carried out by MALDI-TOF/TOF. Results: The SDS-PAGE pattern of LYS treated with papain under nonreducing conditions showed the presence of intact LYS that partially disappeared following reduction with β-mercaptoethanol, releasing IgE-reactive fragments as determined by Western blotting. MALDI-TOF/TOF revealed that papain degraded LYS, giving rise to three IgE-binding fragments: LYS (22-129), LYS (34-96) and LYS (62-128) that likely remained linked through the disulfide bonds present in the LYS molecule. Conclusion: The combined administration of LYS with proteolytic enzymes such as papain may have developed a severe allergic reaction in the patient studied, underlining the importance of considering all the components and their interactions when drugs are to be consumed by allergic persons.

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“…1 Egg white (EW) is assumed as the main culprit with four allergens (ovomucoid-OVM, Gal d 1; ovalbumin-OVA, Gal d 2; ovotransferrin, Gal d 3; and lysozyme, Gal d 4), and egg yolk (EY) comprises two allergens, α-livetin (Gal d 5) and YGP42 (Gal d 6). A few other proteins from EW, EY, or both were reported as IgE-reactive in separate studies, [2][3][4][5][6][7] making it difficult to deduce their relative involvement.…”

Section: Introductionmentioning

confidence: 99%

Relative reactivity to egg white and yolk or change upon heating as markers for baked egg tolerance

Brossard

Rancé

Drouet

et al. 2019

Pediatric Allergy Immunology

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Background Hen's egg food allergy is frequent in childhood and phenotypically heterogeneous. Some children can tolerate extensively heated egg. We investigated whether individual relative responses could differentiate children who tolerate baked egg. Methods Reactivities to raw, pasteurized or hard‐boiled egg (E), egg white (EW), and egg yolk (EY) fractions were tested by skin prick test (SPT) in 54 egg‐allergic children. IgE‐sensitization to EW and EY was determined by ImmunoCAP and IgE‐binding to EW and 8 EW proteins and to EY and 4 EY sub‐fractions by ELISA. Population heterogeneity was assessed by hierarchical ascending classification upon individual variations of reactivity and links between classifications and clinical features by analyzing the contingency tables. Results All children had positive SPT to raw E and raw EW and 72% to raw EY. Heating decreased SPT‐reactivity for some children, pasteurization being less effective than hard‐boiling. Children were classed into three classes from relative SPT‐reactivity to raw fractions, two from variations of SPT‐reactivity with each thermal processing or EW/EY ratio of sensitization, and four from their sensitization pattern. Classifications according to heating were found independent of each other. SPT variations with hard‐boiling, IgE‐sensitization (ratio or pattern) were linked to allowance by the physicians of egg in baked products. Conclusions Egg‐allergic children were often both sensitized to EY and EW, and heterogeneous patterns of relative responses were evidenced. Irrespective of age and level of sensitization, a low EW/EY ratio or SPT getting null with hard‐boiling was found in children allowed to eat baked egg.

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Identification of an IgE Reactive Peptide in Hen Egg Riboflavin Binding Protein Subjected to Simulated Gastrointestinal Digestion

Cited by 9 publications

References 23 publications

Immunological behavior of in vitro digested egg‐white lysozyme

Immunological behavior of in vitro digested egg‐white lysozyme

Anaphylaxis Induced by a Drug Containing Lysozyme and Papain: Influence of Papain on the IgE Response

Relative reactivity to egg white and yolk or change upon heating as markers for baked egg tolerance

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