| Title: | Recombinant glycoproteins resembling carbohydrate-specific IgE epitopes from plants, venoms and mites |
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| Authors: | ID Gattinger, Pia (Author)
ID Mittermann, Irene (Author)
ID Lupinek, Christian (Author)
ID Hofer, Gerhard (Author)
ID Keller, Walter (Author)
ID Bidovec, Urška, Klinika Golnik (Author)
ID Korošec, Peter, Klinika Golnik (Author)
ID Koessler, Christine (Author)
ID Novak, Natalija (Author)
ID Valenta, Rudolf (Author) |
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| Files: |  PDF - Presentation file, download (2,07 MB)
MD5: 877B01939227A3C0DCDACEDEC5C4A6F9
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MD5: 2B96CA34BD1413ED682EF0F9160613E8
 URL - Source URL, visit https://www.ebiomedicine.com/article/S2352-3964(18)30573-5/fulltext
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| Language: | English |
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| Typology: | 1.01 - Original Scientific Article |
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| Organization: |  UKPBAG - University Clinic of Respiratory and Allergic Diseases Golnik
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| Abstract: | Background: N-linked glycans present in venoms, pollen and mites are recognized by IgE antibodies from >20% of allergic patients but have low or no allergenic activity. Objectives: To engineer recombinant glycoproteins resembling carbohydrate-specific IgE epitopes from venoms, pollen and mites which can discriminate carbohydrate-specific IgE from allergenic, peptide-specific IgE. Methods: One or two N-glycosylation sites were engineered into the N-terminus of the non-allergenic protein horse heart myoglobin (HHM) using synthetic gene technology. HHM 1 and HHM 2 containing one or two N-glycosylation sites were expressed in baculovirus-infected High-FiveTM insect cells and a non-glycosylated version (HHM 0) was obtained by mutating the glycosylation motif. Recombinant HHM proteins were analyzed regarding fold and aggregation by circular dichroism and gel filtration, respectively. IgE reactivity was assessed by ELISA, immunoblotting and quantitative ImmunoCAP measurements. IgE inhibition assays were performed to study cross-reactivity with venom, plant and mite-derived carbohydrate IgE epitopes. Results: HHM-glycovariants were expressed and purified from insect cells as monomeric and folded proteins. The HHM-glycovariants exhibited strictly carbohydrate-specific IgE reactivity, designed to quantify carbohydrate specific IgE and resembled IgE epitopes of pollen, venom and mite-derived carbohydrates. IgE-reactivity and inhibition experiments established a hierarchy of plant glcyoallergens (nPhl p 4 > nCyn d 1 > nPla a 2 > nJug r 2 > nCup a 1 > nCry j 1) indicating a hitherto unknown heterogeneity of carbohydrate IgE epitopes in plants which were completely represented by HHM 2. Conclusion: Defined recombinant HHM-glycoproteins resembling carbohydrate-specific IgE epitopes from plants, venoms and mites were engineered which made it possible to discriminate carbohydrate- from peptide-specific IgE reactivity. |
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| Keywords: | allergy and immunology, allergens, glycoproteins, molecular diagnostic technique, recombinant glycoproteins, molecular allergology, component-resolved diagnosis |
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| Publication status: | Published |
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| Publication version: | Version of Record |
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| Place of publishing: | Nizozemska |
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| Publisher: | Elsevier |
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| Year of publishing: | 2019 |
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| Number of pages: | str. 33-43 |
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| Numbering: | Vol. 39 |
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| PID: | 20.500.12556/DiRROS-12589  |
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| UDC: | 616-097 |
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| ISSN on article: | 2352-3964 |
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| DOI: | 10.1016/j.ebiom.2018.12.002 |
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| COBISS.SI-ID: | 2048486001 |
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| Copyright: | © 2018 The Authors |
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| Note: | Soavtorji: Irene Mittermann, Christian Lupinek, Gerhard Hofer, Walter Keller, Urska Bidovec Stojkovic, Peter Korosec, Christine Koessler, Natalija Novak, Rudolf Valenta;
Nasl. z nasl. zaslona;
Opis vira z dne 25. 4. 2019;
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| Publication date in DiRROS: | 16.10.2020 |
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| Views: | 2004 |
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| Downloads: | 1416 |
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