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Reducing Allergenicity to Arginine Kinase from Mud Crab Using Site-Directed Mutagenesis and Peptide Aptamers
journal contribution
posted on 2019-04-09, 00:00 authored by Xue-Jiao Mei, Meng-Si Li, Yang Yang, Meng Liu, Hai-Yan Mao, Ming-Li Zhang, Min-Jie Cao, Guang-Ming LiuThe
mud crab (Scylla paramamosain) is widely
consumed but can cause a severe food allergic reaction. To reduce
allergenicity to arginine kinase (AK), site-directed mutagenesis was
used to destroy disulfide bonds or mutate critical amino acids of
conformational epitopes. Three hypoallergenic mutant AKs (mAK1, mAK2,
and mAK3) were generated, with the immunoreactivity decreasing by
54.2, 40.1, and 71.4%, respectively. In comparison to recombinant
AK (rAK), the structure of mAKs was clearly changed. Additionally,
antisense peptides were designed on the basis of linear epitopes and
pepsin-cutting sites of AK. Five peptide aptamers were screened by
molecular docking and then analyzed by the immunoglobulin E inhibition
enzyme-linked immunosorbent assay and human Laboratory of Allergic
Diseases 2 mast cell degranulation assay. The peptide aptamers could
significantly inhibit allergenicity of rAK and mAKs, and the inhibitory
effect of peptide aptamer 3 was slightly better than the others. These
results provide synergistic methods to reduce allergenicity to AK,
which could be applied to other shellfish allergens.