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Evaluation of cross‐reactivity between casein components using inhibition assay and in silico analysis
Author(s) -
Naito Michihiro,
Matsui Teruaki,
Yamada Chikako,
Tagami Kazunori,
Ito Komei,
Izumi Hidehiko
Publication year - 2021
Publication title -
pediatric allergy and immunology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.269
H-Index - 89
eISSN - 1399-3038
pISSN - 0905-6157
DOI - 10.1111/pai.13405
Subject(s) - in silico , casein , cross reactivity , epitope , peptide , milk allergy , peptide sequence , immunoglobulin e , microbiology and biotechnology , medicine , biochemistry , chemistry , antibody , biology , immunology , cross reactions , gene
Background We previously reported that the specific IgE levels to αs1‐casein (CN) and β‐CN in patients with cow's milk allergy decreased with similar dynamics during oral immunotherapy. Therefore, we hypothesized that αs1‐ and β‐CN have strong cross‐reactivity among CN components, despite the low similarity in the full‐length amino acid sequences. Methods The αs1‐, β‐, and κ‐CN were purified from commercial cow's milk. We recruited 39 patients with cow's milk allergy, and the serum IgE levels for each CN component were measured by enzyme‐linked immunosorbent assay (ELISA). Cross‐reactivity between CN components was investigated by competitive ELISA against αs1‐CN. Sequence homology between CN components at the peptide level was calculated using in silico analysis and quantified by the property distance ( PD ) value. Results The αs1‐CN–specific IgE levels exhibited a strong positive correlation with the β‐CN–specific IgE ( r = 0.945, P < .001). Complete competition was observed by β‐CN against αs1‐CN, suggesting the presence of common epitopes between them. In silico analysis detected 24 peptide sets with PD values lower than 10 between αs1‐ and β‐CN, and 14 sets between αs1‐ and κ‐CN. The amino acid sequences of αs1‐CN (E61‐E70) and β‐CN (I12‐E21) that showed the lowest PD value (5.30) were present in the characteristic sequence known as casein phosphopeptide (CPP). Conclusion We detected strong cross‐reactivity between CN components. Furthermore, we found highly homologous sequences in the CPP region, which contains a core sequence of "SSSEE" with phosphorylated serine residues.