A hypo-immunoreactive derivative lacking the dominant linear Scylla
paramamosain allergen epitope
Abstract
Background: Scylla paramamosain frequently elicits IgE-mediated type-I
hypersensitivity reactions. Molecular candidates for crab
allergen-specific immunotherapy (AIT) have not been studied previously.
This study aimed to investigate the effects of conformational and linear
epitopes on immunoreactivity, and to produce hypo-immunoreactive
derivatives for crab myofibril allergens. Methods: We produced reduced
and alkylated allergens (alkylation performed using iodoacetamide
[IAA]) with destroyed conformational epitopes. We also produced
mutant (mt) allergens with deleted or heat/digestion-treated linear
epitopes. These allergens are collectively referred to here as
derivatives. Structural changes in the derivatives were determined using
circular dichroism and fluorescent probes, and the immunoreactivity of
derivatives was analyzed by performing enzyme-linked immunosorbent
assays (ELISAs) and basophil-activation tests (BATs). Results: Compared
with wild-type allergens (wtALLERGENs), IAA-treated allergens had
dramatically altered protein structures, whereas mutant allergens
(mtALLERGENs) showed slight structural effects. ELISAs revealed the
heterogeneous epitope-recognition patterns with derivatives among 29
crab-sensitive patients, of whom 13% and 62% recognized conformational
and linear epitopes, respectively, whereas 25% recognized both epitope
types to the same extent. mtALLERGENs had lower immunoreactivity than
IAA-treated wtALLERGENs, showing no intergroup difference versus
IAA-treated mtALLERGENs. Furthermore, mtALLERGENs could not bind to IgE
or induce basophil activation in some patients. Conclusions:
Heat/digestion treatment of linear epitopes had a greater influence on
immunoreactivity than their structural destruction. Conformational and
linear epitopes were recognized in patient-specific crab myofibril
allergens. Our results imply that hypo-immunoreactive derivatives of
crab myofibril allergens that specifically lacked linear epitopes may
serve as viable candidates for AIT in patients with crab allergies.