Immunochemical and physical properties of peptic-digested ovomucoid

Matsuda, Tsukasa; Watanabe, Kenji; Nakamura, Ryo

doi:10.1021/jf00119a005

Cited by 42 publications

(38 citation statements)

References 16 publications

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“…OVM accounts for about 11% of all egg white proteins [6]and has a molecular weight of 28 kDa, containing a carbohydrate content of 20–25% [7]. OVM is known to be stable to digestion and heat, and cooked eggs can cause allergic reactions in OVM-specific allergic patients [8, 9, 10, 11]. One possible reason for this is that OVM contains linear epitopes that are only slightly affected by conformational changes induced by heat denaturation.…”

Section: Introductionmentioning

confidence: 99%

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Kinetic Analysis of Pepsin Digestion of Chicken Egg White Ovomucoid and Allergenic Potential of Pepsin Fragments

Takagi

Teshima

Okunuki

et al. 2005

Int Arch Allergy Immunol

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Background: The allergenic potential of chicken egg white ovomucoid (OVM) is thought to depend on its stability to heat treatment and digestion. Pepsin-digested fragments have been speculated to continue to exert an allergenic potential. OVM was digested in simulated gastric fluid (SGF) to examine the reactivity of the resulting fragments to IgE in sera from allergic patients. Methods: OVM was digested in SGF and subjected to SDS-PAGE. The detected fragments were then subjected to N-terminal sequencing and liquid chromatography/mass spectrometry/mass spectrometry analysis to confirm the cleavage sites and partial amino acid sequences. The reactivity of the fragments to IgE antibodies in serum samples from patients allergic to egg white was then determined using Western blotting (n = 24). Results: The rate of OVM digestion depended on the pepsin/OVM ratio in the SGF. OVM was first cleaved near the end of the first domain, and the resulting fragments were then further digested into smaller fragments. In the Western blot analysis, 93% of the OVM-reactive sera also bound to the 23.5- to 28.5-kDa fragments, and 21% reacted with the smaller 7- and 4.5-kDa fragments. Conclusion: When the digestion of OVM in SGF was kinetically analyzed, 21% of the examined patients retained their IgE-binding capacity to the small 4.5-kDa fragment. Patients with a positive reaction to this small peptide fragment were thought to be unlikely to outgrow their egg white allergy. The combination of SGF-digestibility studies and human IgE-binding experiments seems to be useful for the elucidation and diagnosis of the allergenic potential of OVM.

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Section: Introductionmentioning

confidence: 99%

“…We previously reported the digestibility of 10 kinds of food proteins in simulated gastric fluid (SGF) [8, 14]. OVM was digested relatively rapidly, but several fragments were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) followed by Coomassie blue (CBB) staining.…”

Section: Introductionmentioning

confidence: 99%

Kinetic Analysis of Pepsin Digestion of Chicken Egg White Ovomucoid and Allergenic Potential of Pepsin Fragments

Takagi

Teshima

Okunuki

et al. 2005

Int Arch Allergy Immunol

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“…The titer of IgE antibodies to ovomucoid was significantly greater in the patients with positive results to oral egg challenge tests compared to the patients with negative results. This may result from the unique characteristics of ovomucoid such as relative stability against digestion with proteinases [12]and heat [13]as well as strong allergenicity [9, 10]compared with other egg white components. However, there was a large extent of overlapping of IgE antibody titers for ovomucoid between both groups.…”

Section: Introductionmentioning

confidence: 99%

“…However, there was a large extent of overlapping of IgE antibody titers for ovomucoid between both groups. According to Matsuda et al [12], this allergenicity and antigenicity were still retained after fragmentation by pepsin digestion.…”

Section: Introductionmentioning

confidence: 99%

Clinical Significance of IgE–Binding Activity to Enzymatic Digests of Ovomucoid in the Diagnosis and the Prediction of the Outgrowing of Egg White Hypersensitivity

Urisu

Yamada

Tokuda

et al. 1999

Int Arch Allergy Immunol

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Background: We frequently encounter subjects without overt symptoms despite high IgE antibodies to egg white and its components. The measurements of these antibodies are not necessarily efficient for the diagnosis or the prediction of the outcome of egg allergy in children. Methods: Specific IgE antibodies to egg white and its components, including ovomucoid, ovalbumin, ovotransferrin and lysozyme, were measured by direct RAST assays. IgE–binding activity to ovomucoid degraded by pepsin, trypsin and chymotrypsin was examined by RAST inhibition. Thirty subjects were divided into two groups with positive (n=18; mean age ± SD = 42 ±25 months) and negative (n=12; mean age ± SD = 48 ±31 months) oral challenge tests with egg white antigens. The individuals with positive results to the first challenge tests were given the second provocation tests at mean intervals of 32 months. IgE–binding activity of the sera collected on the first challenge to these ovomucoid fragments was compared between subjects with positive and negative reactions to the follow–up challenge tests. Results: There were no significant differences in IgE antibody titers to egg white and its components between the positive and negative groups at the first and the second challenge tests. IgE–binding activity to ovomucoid digests after treatments with pepsin (p = 0.000008) and trypsin (p=0.037), except chymotrypsin (p=0.062), were significantly higher in subjects with positive challenge tests than in those with negative results. The difference was most remarkable in the IgE–binding to pepsin digests; the average concentrations (mean – SD and mean + SD) needed for 50% RAST inhibition in the positive group and in the negative group were 2.6 μg/ml (0.3 and 25) and 94.2 μg/ml (24.7 and 358.7), respectively. A significant difference was still observed in the inhibition tests using filtrates of pepsin digests with a membrane with MW 10,000 (p=0.014) and 3,000 (p=0.042) of cutoff. The concentration (mean= 0.8, mean – SD=0.2, mean + SD=3.4; μg/ml) of pepsin–treated ovomucoid required for 50% RAST inhibition in the subjects with positive second challenge results was significantly (p=0.033) lower than that (mean=6.8, mean–SD=0.6, mean + SD=73.9) of the negative group. Conclusion: IgE–binding activity to pepsin–digested ovomucoid was of diagnostic value to distinguish the challenge–positive subjects from the negative subjects. Subjects with high IgE–binding activity to pepsin–treated ovomucoid are unlikely to outgrow egg white allergy.

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“…Although each peptide fragment with molecular weight of 22.0, 17.8, or 17.0kDa showed broad bands on SDS-PAGE, they gave the same sequences for the N-terminal side. It is known that peptide fragments possessing carbohydrate chains give a broad migration band on SDS-PAGE (19). Accordingly, the molecular size of each fragment may depend on the difference in sequence of the C-terminal side.…”

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confidence: 99%

Immunochemical Characterization of Ovomucoid from Japanese Quail Egg White Using Monoclonal Antibodies.

Takahashi

Horiguchi

Bando

et al. 1999

Journal of Nutritional Science and Vitaminology, J Nutr Sci Vit

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SummaryThe ovomucoid of Japanese quail egg white is known as a proteinase inhibitor or protein component responsible for egg allergy. In order to characterize the antigenic properties of ovomucoid in relation to its molecular structure, we have prepared two monoclonal antibodies, E9 (IgG1) and E11 (IgG1), recognizing distinct epitopes from each other. These monoclonal antibodies bound to the SDS/mercaptoethanol-treated ovomucoid, but not to the reductively carboxymethylated and pyridyl ethylated ovomucoid. By immunoblotting analysis of the peptic digests of ovomucoid, it was shown that E9 bound to the fragment consisting of two domains, I and II, of the ovomucoid, and that E11 reacted with the fragment containing domain III. These results indicate that the antigenic activity depends on the conformational structure of domains tightly folded by disulfide linkages. Ovomucoids from hen and duck were also recognized by both the antibodies, although having less affinity compared to the one from Japanese quail. These antibodies proved to be effective in establishing an enzyme-linked immunosorbent assay system for quantitative analysis of quail ovomucoid.

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Immunochemical and physical properties of peptic-digested ovomucoid

Cited by 42 publications

References 16 publications

Kinetic Analysis of Pepsin Digestion of Chicken Egg White Ovomucoid and Allergenic Potential of Pepsin Fragments

Kinetic Analysis of Pepsin Digestion of Chicken Egg White Ovomucoid and Allergenic Potential of Pepsin Fragments

Clinical Significance of IgE–Binding Activity to Enzymatic Digests of Ovomucoid in the Diagnosis and the Prediction of the Outgrowing of Egg White Hypersensitivity

Immunochemical Characterization of Ovomucoid from Japanese Quail Egg White Using Monoclonal Antibodies.

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