Structural analysis of 8-anilino-1-naphthalene sulfonate (ANS) binding to the PR-10 allergen Ara h 8

Abstract

We previously determined crystal structures of peanut allergen Ara h 8.0101 in the apo form as well as in complex with model ligands. These structures illustrated the varied ligand binding capabilities of PR-10s and Ara h 8's structural similarity to the major birch allergen Bet v 1. Here, we expanded on those structural studies with structures of Ara h 8.0101 and Ara h 8.0201 in complex with 8-anilino-1-naphthalene sulfonate (ANS), as well as the apo form of Ara h 8.0201. Structural studies revealed that both proteins may bind more than one ANS molecule. We also examined the impact of ANS on the ligand binding cavities of Ara h 8.0101 and Ara h 8.0201 with fluorescence assays and compared the results to prototypic PR-10 Bet v 1.0101. Moreover, as ANS is often used in fluorescence-based ligand binding assays, we analyzed structures from the PDB and provided a summary on experimentally determined ANS binding sites. These analyses show that ANS is useful for investigation of ligand binding sites, but it may also participate in non-specific reactions on nonpolar surfaces of proteins.

Keywords: ANS; Allergen; Crystal structure; Ligand-binding; PR-10; Peanut.