Analysis of the Structural and Immunological Stability of 2S Albumin, Nonspecific Lipid Transfer Protein, and Profilin Allergens from Mustard Seeds

Departamento de Bioquímica y Biología Molecular, Facultad de Química, Universidad Complutense de Madrid, Ciudad Universitaria, 28040 Madrid, Spain.
Journal of Agricultural and Food Chemistry (Impact Factor: 2.91). 05/2012; 60(23). DOI: 10.1021/jf300555h
Source: PubMed

ABSTRACT This work investigates the resistance to proteolysis and heating of the yellow mustard (Sinapis alba L.) allergens Sin a 1 (2S albumin), Sin a 3 (nonspecific lipid transfer protein, LTP), and Sin a 4 (profilin) to explain their potential capability to induce primary sensitization at the gastrointestinal level. Sin a 1 and Sin a 3 resisted gastric digestion showing no reduction of the IgE reactivity. Intestinal digestion of Sin a 1 and Sin a 3 produced a limited proteolysis but retained significant IgE-binding reactivity. Sin a 1 was stable after heating, and although Sin a 3 was modified, most of its structure was recovered after cooling back. These two allergens would be therefore able to sensitize by ingestion. Sin a 4 was completely digested by gastric treatment and its conformational structure markedly modified at 85 °C. Thus, this allergen can be described as a nonsensitizing mustard allergen.

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    • "In accordance with this finding, CD measurement of Cor a 14 revealed that the protein retained most of its secondary structure after heating and completely refolded after cooling. This was also shown for other 2S albumins [37] [38]. "
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    ABSTRACT: Allergens from nuts frequently induce severe allergic reactions in sensitive individuals. The aim of this study was to elucidate the physicochemical characteristics of natural Cor a 14, the 2S albumin from hazelnut. Cor a 14 was purified from raw hazelnuts using a combination of precipitation and chromatographic techniques. The protein was analyzed using gel electrophoresis, mass spectrometry, and far-UV circular dichroism (CD) analyses. The IgE binding of native, heat-treated, and in vitro digested Cor a 14 was studied. We identified two different Cor a 14-isoforms and showed microclipping at the C-terminus. CD spectra at room temperature showed the typical characteristics of 2S albumins, and temperatures of more than 80°C were required to start unfolding of Cor a 14 demonstrating its high stability to heat treatment. In vitro digestion experiments revealed that Cor a 14 is resistant to proteolytic degradation. Native and heat-treated protein was recognized by sera from hazelnut allergic patients. However, denaturation of the allergen led to significantly reduced IgE binding. We identified two different isoforms of Cor a 14 displaying high stability under heating and gastric and duodenal conditions. Data from IgE binding experiments reveal the existence of both, linear and conformational epitopes. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved.
    Molecular Nutrition & Food Research 07/2015; DOI:10.1002/mnfr.201500071 · 4.60 Impact Factor
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    • "Four allergens from yellow mustard seeds have been identified, purified and characterized so far: i) 2S albumin Sin a 1 (14 kDa) [10,11]; ii) 11S globulin Sin a 2 (51 kDa) [12,13]; iii) LTP Sin a 3 (12 kDa) [14]; and iv) profilin Sin a 4 (13–14 kDa) [14]. Sin a 1 and Sin a 3 but not Sin a 4 might act as genuine food allergens able to reach the gut immune system due to their high structural and immunological stability [15]. The capacity of Sin a 2 to act as primary sensitizer at the intestinal mucosa has not been investigated so far. "
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    ABSTRACT: Background The 11S globulin Sin a 2 is a marker to predict severity of symptoms in mustard allergic patients. The potential implication of Sin a 2 in cross-reactivity with tree nuts and peanut has not been investigated so far. In this work, we studied at the IgG and IgE level the involvement of the 11S globulin Sin a 2 in cross-reactivity among mustard, tree nuts and peanut. Methods Eleven well-characterized mustard-allergic patients sensitized to Sin a 2 were included in the study. A specific anti-Sin a 2 serum was obtained in rabbit. Skin prick tests (SPT), enzyme-linked immunosorbent assay (ELISA), immunoblotting and IgG or IgE-inhibition immunoblotting experiments using purified Sin a 2, Sin a 1, Sin a 3, mustard, almond, hazelnut, pistachio, walnut or peanut extracts were performed. Results The rabbit anti-Sin a 2 serum showed high affinity and specificity to Sin a 2, which allowed us to demonstrate that Sin a 2 shares IgG epitopes with allergenic 11S globulins from tree nuts (almond, hazelnut, pistachio and walnut) but not from peanut. All the patients included in the study had positive skin prick test to tree nuts and/or peanut and we subdivided them into two different groups according to their clinical symptoms after ingestion of such allergenic sources. We showed that 11S globulins contain conserved IgE epitopes involved in cross-reactivity among mustard, tree nuts and peanut as well as species-specific IgE epitopes. Conclusions The allergenic 11S globulin Sin a 2 from mustard is involved in cross-reactivity at the IgE level with tree nuts and peanut. Although the clinical relevance of the cross-reactive IgE epitopes present in 11S globulins needs to be investigated in further detail, our results contribute to improve the diagnosis and management of mustard allergic patients sensitized to Sin a 2.
    12/2012; 2(1):23. DOI:10.1186/2045-7022-2-23
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    ABSTRACT: In this study, we used a mass spectrometry-based quantification approach employing isotopic (ICPL) and isobaric (iTRAQ) labeling to investigate the pattern of protein deposition during castor oil seed (Ricinus communis L.) development, including that of proteins involved in fatty acid metabolism, seed storage proteins, toxins and allergens. Additionally, we have used an off-line Hydrophilic Interaction Chromatography (HILIC) as a step of peptide fractionation preceding the Reverse Phase nanoLC coupled with a LTQ Orbitrap. We were able to identify a total of 1875 proteins and from these 1748 could be mapped to extant castor gene models, expanding considerably the number proteins so far identified from developing castor seeds. Cluster validation and statistical analysis resulted in 975 protein trend patterns and the relative abundance of 618 proteins. The results presented in this work give important insights into certain aspects of the biology of castor oil seed development such as carbon flow, anabolism and catabolism of fatty acid and the pattern of deposition of reserve proteins, toxins and allergens such as ricin and 2S albumins. We also found, for the first time, some genes of reserve proteins that are differentially expressed during seed development.
    Journal of Proteome Research 10/2013; 12(11). DOI:10.1021/pr400685z · 4.25 Impact Factor
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