Characterization and identification of allergen epitopes: recombinant peptide libraries and synthetic, overlapping peptides

Reese, Gerald; Ayuso, Rosalía; Leong-Kee, Susan; Plante, Matthew; Lehrer, Samuel B.

doi:10.1016/s0378-4347(01)00104-9

Cited by 24 publications

(18 citation statements)

References 33 publications

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“…This has been demonstrated in the case of Ara h 1 19 and Ara h 2 from peanuts, 12 b-lactoglobulin from cow's milk, 20 and Pen a 1 from shrimp. 21 The linear epitopes are generally known to be resistant to heat and proteolysis, 22 and they tend to retain their allergenic reactivity, whereas many conformational epitopes of native food proteins are modified or disrupted by heat, chemical treatments, or both, thus losing their IgE-binding potential. 1 Such treatments usually result in exposure of linear epitopes as well.…”

Section: Discussionmentioning

confidence: 99%

Identification and characterization of linear B-cell epitopes of β-globulin, a major allergen of sesame seeds

Wolff¹,

Yannai²,

Karin

et al. 2004

Journal of Allergy and Clinical Immunology

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Section: Discussionmentioning

confidence: 99%

Identification and characterization of linear B-cell epitopes of β-globulin, a major allergen of sesame seeds

Wolff¹,

Yannai²,

Karin

et al. 2004

Journal of Allergy and Clinical Immunology

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“…At present the SPOTs system (Genosys, The Woodland, Tex.) and the Novatope system (Novagen, Madison, Wis.) are extensively used to identify IgE-binding epitopes, and the results obtained with the two systems have been extensively compared (25). Moreover, fragmented peptides are reported to have higher IgE-binding capacities than whole molecules in the case of paramyosin, Der f 11 (33).…”

Section: Discussionmentioning

confidence: 99%

Analysis of Amino Acid Sequence Variations and Immunoglobulin E-Binding Epitopes of German Cockroach Tropomyosin

Jeong

Lee

et al. 2004

Clin Vaccine Immunol

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The allergenicities of tropomyosins from different organisms have been reported to vary. The cDNA encoding German cockroach tropomyosin (Bla g 7) was isolated, expressed, and characterized previously. In the present study, the amino acid sequence variations in German cockroach tropomyosin were analyzed in order to investigate its influence on allergenicity. We also undertook the identification of immunodominant peptides containing immunoglobulin E (IgE) epitopes which may facilitate the development of diagnostic and immunotherapeutic strategies based on the recombinant proteins. Two-dimensional gel electrophoresis and immunoblot analysis with mouse anti-recombinant German cockroach tropomyosin serum was performed to investigate the isoforms at the protein level. Reverse transcriptase PCR (RT-PCR) was applied to examine the sequence diversity. Eleven different variants of the deduced amino acid sequences were identified by RT-PCR. German cockroach tropomyosin has only minor sequence variations that did not seem to affect its allergenicity significantly. These results support the molecular basis underlying the cross-reactivities of arthropod tropomyosins. Recombinant fragments were also generated by PCR, and IgE-binding epitopes were assessed by enzyme-linked immunosorbent assay. Sera from seven patients revealed heterogeneous IgE-binding responses. This study demonstrates multiple IgE-binding epitope regions in a single molecule, suggesting that full-length tropomyosin should be used for the development of diagnostic and therapeutic reagents.

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“…To verify the allergenicity of the released peptides, a dotblotting protocol [20] was performed on the purified proteolytic peptides (details of the purification in Section 2). After spotting the different purified products on the PVDF membrane, incubation with allergic patient serum was performed.…”

Section: Allergenicity Of the Purified Peptides Derived From Simulatementioning

confidence: 99%

In vitro gastrointestinal digestion of the major peach allergen Pru p 3, a lipid transfer protein: Molecular characterization of the products and assessment of their IgE binding abilities

Cavatorta

Sforza

Aquino

et al. 2010

Molecular Nutrition Food Res

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A simulated gastrointestinal digestion has been carried out on purified peach lipid transfer protein, one of the main allergens among the population of the Mediterranean area and the major allergen of peach allergic patients. The percentage of intact protein, after extensive digestion, measured by comparison with a non-digestible peptide analogue used as internal standard, was found to be about one-third of the original protein content. The peptides formed in digested fraction were characterized by means of LC/MS. The products of the digestion essentially derived from trypsin action, whereas the protein appeared to be resistant to pepsin and chymotrypsin. The identified peptides could be classified as low molecular weight and high molecular weight peptides. The latter consisted of the full protein, with the disulfide bridges still intact, deprived of the smaller peptides. The different digestion products, including the high and low molecular weight peptides, were purified by LC and assessed, together with the intact protein, by dot-blot analysis with sera of allergic patients, allowing to estimate their potential allergenicity. The intact protein and the high molecular weight peptides were found to be recognized by patients' sera, whereas the small peptides were found to be not reactive.

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Characterization and identification of allergen epitopes: recombinant peptide libraries and synthetic, overlapping peptides

Cited by 24 publications

References 33 publications

Identification and characterization of linear B-cell epitopes of β-globulin, a major allergen of sesame seeds

Identification and characterization of linear B-cell epitopes of β-globulin, a major allergen of sesame seeds

Analysis of Amino Acid Sequence Variations and Immunoglobulin E-Binding Epitopes of German Cockroach Tropomyosin

In vitro gastrointestinal digestion of the major peach allergen Pru p 3, a lipid transfer protein: Molecular characterization of the products and assessment of their IgE binding abilities

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