Mapping of IgE epitopes in in vitro gastroduodenal digests of β-lactoglobulin produced with human and simulated fluids
This work aimed to identify the IgE epitopes of the allergenic whey protein β-lactoglobulin (β-LG) following in vitro gastrointestinal digestion using human (HF) or simulated digestive fluids (SF). The IgE-binding of the digests was evaluated by inhibition ELISA with sera from milk allergic patients and the low molecular weight digestion products were identified by RP-HPLC-MS/MS. These peptides were then chemically synthesized and analyzed by a peptide microarray immunoassay. β-LG resisted digestion during the gastric phase, while no residual β-LG was observed at the end of the duodenal phase. The immunoreactivity of the digests was consistent with the levels of intact β-LG present, showing almost negligible IgE binding after gastrointestinal digestion with both systems. There were similarities in the peptide patterns produced, showing, respectively, 21 and 18 identical peptides in the gastric and duodenal digests. Digestion products related to five high-frequency IgE-binding synthetic peptides: β-LG (43-60), β-LG (47-62), β-LG (86-99), β-LG (86-100) and β-LG (135-147), which were released upon in vitro digestion, particularly with SF. These matched two very immunoreactive areas of the protein: 43-68 and 122-146, as mapped using a library of 36 20-amino acid peptides corresponding to the primary sequence of β-LG. However, no IgE-binding peptides comprising the residues 86-100 were detected by analyzing the 36-peptide library by microarray immunoassay. © 2014.
| Main Authors: | , , , , |
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| Other Authors: | |
| Format: | artículo biblioteca |
| Published: |
Elsevier
2014
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| Subjects: | Milk allergy, β-Lactoglobulin, In vitro digestion, Human digestive fluids, IgE-binding, Peptide microarray immunoassay, |
| Online Access: | http://hdl.handle.net/10261/113588 http://dx.doi.org/10.13039/501100003329 http://dx.doi.org/10.13039/501100003339 http://dx.doi.org/10.13039/501100000780 http://dx.doi.org/10.13039/100006423 |
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