Reconstruction of fish allergenicity from the content and structural traits of the component β-parvalbumin isoforms

Abstract

Most fish-allergic patients have anti-β-parvalbumin (β-PV) immunoglobulin E (IgE), which cross-reacts among fish species with variable clinical effects. Although the β-PV load is considered a determinant for allergenicity, fish species express distinct β-PV isoforms with unknown pathogenic contributions. To identify the role various parameters play in allergenicity, we have taken Gadus morhua and Scomber japonicus models, determined their β-PV isoform composition and analyzed the interaction of the IgE from fish-allergic patient sera with these different conformations. We found that each fish species contains a major and a minor isoform, with the total PV content four times higher in Gadus morhua than in Scomber japonicus. The isoforms showing the best IgE recognition displayed protease-sensitive globular folds, and if forming amyloids, they were not immunoreactive. Of the isoforms displaying stable globular folds, one was not recognized by IgE under any of the conditions, and the other formed highly immunoreactive amyloids. The results showed that Gadus morhua muscles are equipped with an isoform combination and content that ensures the IgE recognition of all PV folds, whereas the allergenic load of Scomber japonicus is under the control of proteolysis. We conclude that the consideration of isoform properties and content may improve the explanation of fish species allergenicity differences.

Animal Food Allergens Share the Formation of IgE-binding Amyloids

“…In-silico analysis using previously described IgE-binding epitopes and the amyloid forming segments predicted by the ZipperDB algorithm shows that each allergen chain contains multiple regions with a functional overlap. This behaviour was described for the segments 25 FDHKAFFTKVGLAAKSSA 42 and 67 FLQNFSAGARAL 78 of Gad m 1 using anti-fibril OC-antibody and sera IgE interactions [15,16]. Interestingly, the N-terminal region of each of the segments (DHKAFFTKV and FLQNFS) form part of helical structures in the native state of Gad m 1 [26].…”

“…This conformational multiplicity of the regions forming the IgE-binding epitopes questions the validity of mapping the epitopes singly on the native 3D structures of food allergens. In fact, refolding and amyloid assembly of the IgE-binding epitopes will impede their hydrolysis by proteases and will amplify the avidity and affinity of any ligand binding process [12,14,15,25]. On the other hand, the identified segments significantly differ in the sequence which allows specificity in the IgE binding.…”

Proteomics-Based Methodologies for the Detection and Quantification of Seafood Allergens

“…Moreover, the complete sequence of four β-PRVB isoforms (PRVB1 (P86431), PRVB1.1, PRVB2 (P86432), and PRVB2.1 variants) from farmed rainbow trout (Oncorhynchus mykiss) were achieved using matrix-assisted laser desorption (MALDI)MS and MS/MS analysis [53]. Such an isoform diversity impacts the stabilization of β-PRVBs as amyloids under gastric-like conditions, and consequently, their resistance to proteases and IgE binding intensity [54][55][56]. In agreement with that, a shotgun proteomics analysis of 15 different fish species, the protein-based bioinformatics analysis and IgE reactive approaches, was used to identify a total of 35 peptides as B-cell epitopes for all the β-PRVBs included in the UniProtKB database [57] An easy and robust method for fish allergen detection has been developed by utilizing the high-speed, high-resolution, and fragmentation capabilities of the Orbitrap Fusion mass spectrometer implemented with an ultraviolet photodissociation (UVPD) source.…”

“…It must be recalled that extracts are usually prepared from raw fish muscles, limiting the presence of collagen that is more abundant in skin and the effects of processing. Furthermore, fish species muscles vary in the number and relative amount of the expressed β-PRVBs, each of which differs in stability and IgE interaction, introducing variables such as isoallergen composition, temperature treatments, and protease content, which is not yet controlled [56]. Indeed, about 30% of the individuals of a patient cohort having cross-reactive anti-β-PRVB IgEs showed oral tolerance to at least one of the fish species tested [94].…”

Distinct Animal Food Allergens Form IgE-Binding Amyloids

“…Gal d 2 (A-2512), Bos d 5 (L3908) and Bos d 12 (C0406) were purchased from Sigma-Aldrich. Gad m 1 (A5I874) was prepared as described [31]. Before their use all proteins were extensively dialyzed at 4 • C against either 25 mM Tris, 0.1 M NaCl pH 7.5 or 0.1 M Gly pH 1.5 using dialysis membranes with an 8 kDa pore diameter (Spectra Por).…”