β-parvalbumins represent one of the largest animal food-allergen families and are considered cross-reactive pan-allergens in fish. The β-parvalbumin structure is characterized by the presence of three EF-hand motifs (helix-loop-helix) called AB, CD and EF, but only CD and EF can chelate calcium ions.
In order to contribute to the understanding of the allergenicity and the importance of the Ca2+-binding motifs for the stability of β-parvalbumins, we studied the major allergen of cod (Gadus morhua), Gad m 1, a member of the parvalbumin protein family
Methods: The solution structure and the molecular dynamics of Gad m 1 were determined using NMR spectroscopy. Our strategy included high pressure to perturb the system and to evaluate crucial residues for structure stabilization at the atomic level. The Gad m 1-scFv complex structural characterization was done using chemical shift perturbation and the molecular dynamics of the complex was assessed by 15N-relaxation experiments.
Results: Gad m 1 possesses the typical parvalbumin fold that is characterized by the presence of three domains, the two calcium-binding domains CD and EF, and the silent domain AB. High-pressure NMR revealed the important contribution of the AB domain to the protein fold stabilization. Although the Gad m 1 structure and accessibility of putative IgE epitopes are similar to parvalbumins from mackerel and carp, the charge of each of these sites is different.
Conclusions: Our results offer new insights into the design of mutated proteins that would be stable in the non-allergenic apo form. Comparison of the Gad m 1 structure with other parvalbumins was done to understand the observed cross-reactivity.