Negatively charged food additive dye Allura Red rapidly induces SDS-soluble amyloid fibril in beta-lactoglobulin protein

Recent studies have led to an increased interest to categorize small molecular inhibitors of protein fibrillation. In this study, we used spectroscopy, microscopy and gel electrophoresis techniques that provides an elaborated description of the Allura Red-induced amyloid fibrillation in the beta-LG protein at two pHs (7.4 and 3.5). The spectroscopy results show that beta-LG protein form aggregates in the presence of Allura Red (0.04-15.0 mM) at pH 3.5 due to electrostatic and hydrophobic interactions. However, at pH 7.4, the beta-LG does not interact electrostatically with Allura Red and therefore no aggregation occurred. The Allura Red-induced aggregates have an amyloid-like structure that was confirmed by far-UV CD, Congo Red and transmission electron microscopy (TEM). The CD spectrum of beta-LG contains single minima at similar to 218 nm, which shifts towards higher wavelength minima at 225 nm in the presence of Allura Red, characteristics of the cross beta-sheet structure. The TEM results suggest that beta-LG form long straight fibril when exposed to Allura Red at pH 3.5. The Allura Red-induced amyloid fibril is SDS-soluble confirmed by SDS-PAGE techniques. A far UV CD result shows the conversion of Allura Red induced cross beta-sheet structure into alpha-helical structure in the presence of increasing concentration of SDS. The results of this study suggest that the electrostatic, as well as hydrophobic interactions play an important role during Allura Red-induced beta-LG fibrillation. (C) 2017 Published by Elsevier B.V.