Physicochemical characterization and linear epitopes identification of arginine kinase allergen from Crassostrea gigas

Abstract

BACKGROUND Molluscan shellfish, including oysters, often cause allergic reactions in sensitive people throughout the world. It has been demonstrated that arginine kinase (AK) is one of the major allergens of oyster. The present study aimed to evaluate the immunoreactivity and structure of oyster AK as affected by heat treatment, pH change, and in vitro digestion. What is more, the immunoglobulin E‐binding epitopes of this allergen were also predicted and validated. RESULTS Thermal and pH assays revealed that AK was unstable at temperature >40 °C or pH ≤5.0 by sodium dodecyl sulfate polyacrylamide gel electrophoresis and circular dichroism, and the digestibility assays suggested that AK was more easily digested by pepsin than by trypsin and chymotrypsin. The potential epitopes were predicted through immunoinformatics tools, and seven linear epitopes were identified by indirect competition enzyme‐linked immunosorbent assay with pooled sera and individual serum from oyster‐allergic patients. The critical amino acids in each epitope were also confirmed using mutant peptides. These linear epitopes and critical amino acids were apt to distribute on the outer surface of homology‐based AK model. Moreover, the three denaturants (sodium dodecyl sulfate, β‐mercaptoethanol, and urea) can destroy the spatial structure of AK and increase or reduce its allergenicity by denaturation treatments. CONCLUSION Processing conditions lay the foundation for the variation of allergenicity. Seven linear epitopes and their critical amino acids were identified by indirect competitive enzyme‐linked immunosorbent assay. These findings will be helpful in allergy diagnosis and development of hypoallergenic products in the near future. © 2021 Society of Chemical Industry.

Identification of the Immunoglobulin E Epitope of Arginine Kinase, an Important Allergen from Crassostrea angulata

“…Among the linear epitopes identified in this study, L-Cra a 2-2 partially overlapped with the linear epitopes of Crassostrea gigas AK, Scylla paramamosain AK, and L. vannamei AK. − Among the four identified conformational epitopes, C-Cra a 2-4 is partially consistent with conformational epitopes of S. paramamosain AK (I 121 H 185 ­K 189 G 310 T 311 R 312 ­T 316 A 318 E 319 G 320 ­V 322 ), indicating that epitopes of AK are conserved among crustaceans and mollusks .…”

Identification of major and cross‐reactive allergens of local freshwater snail (Pila polita) and the impact of thermal and non‐thermal food processing on allergen stability

“…This result revealed the existence of generally strong cross‐reactivity of the 42 and 49 kDa bands between P. polita and sea snail C. obtusa , whereas moderate cross‐reactivity was observed between P. polita and the other shellfish extracts tested, suggesting the presence of homologous epitopes between P. polita and C. obtusa and the existence of species‐specific epitopes between P. polita and the other shellfish tested. Additionally, the 42 kDa band might consist of several different proteins at the same molecular weight with different allergenic properties, probably arginine kinase, which had been documented as a pan‐allergen in molluscan and crustacean shellfish 19,27,44 . It was interesting to note that the IgE reactivity in one of the sera tested (patient 4) showed an exclusive IgE cross‐reactivity pattern compared with the other sera tested: almost no inhibition reactivity was observed for P. polita against P. monodon , S. olivacea , and L. edulis extracts, but high cross‐reactivity was present with C. obtusa extract, indicating that this patient was highly reactive to only snail allergens, probably due to snail mono‐sensitivity 45…”

“…Additionally, the 42 kDa band might consist of several different proteins at the same molecular weight with different allergenic properties, probably arginine kinase, which had been documented as a pan-allergen in molluscan and crustacean shellfish. 19,27,44 It was interesting to note that the IgE reactivity in one of the sera tested (patient 4) showed an exclusive IgE cross-reactivity pattern compared with the other sera tested: almost no inhibition reactivity was observed for P. polita against P. monodon, S. olivacea, and L. edulis extracts, but high cross-reactivity was present with C. obtusa extract, indicating that this patient was highly reactive to only snail allergens, probably due to snail mono-sensitivity 45 The findings in this study support a previous report from Emoto et al 18 that indicated IgE cross-reactivity is recognized among molluscs, among crustaceans, and between molluscs and crustaceans. The primary structures of crustacean tropomyosin were proved to share more than 90% sequence identity with one another, except for some species showing 60% sequence identity of tropomyosin with other shellfish.…”

“…Cross‐reactivity occurs when an adaptive immune response to a particular antigen causes reactivity to other antigens from other sources that are structurally related to the inducer 14 . Cross‐reactivity among snail allergens has been described between Haliotis rufescens and Haliotis discus discus and other snails such Haliotis diversicolor , Biompahalaria glabrata , Helix aspersa , T. cornutus , and Neptunea polycostata 18,19 . These cross‐reactivity associations were reported due to high frequency of homologous sequence identity in their tropomyosin, between 74 and 100%.…”

“…14 Cross-reactivity among snail allergens has been described between Haliotis rufescens and Haliotis discus discus and other snails such Haliotis diversicolor, Biompahalaria glabrata, Helix aspersa, T. cornutus, and Neptunea polycostata. 18,19 These crossreactivity associations were reported due to high frequency of homologous sequence identity in their tropomyosin, between 74 and 100%. Meanwhile, cross-reactivity also occurs between snails and other molluscs and crustaceans with homologous sequence identity of tropomyosin between 62% and 84%.…”