SUMMARY
The applicability of the novel cyanine dye AK 3-1 to the detection and characterization of pathogenic protein aggregates, amyloid fibrils, was tested using the absorption spectroscopy technique. In an organic solvent dimethyl sulfoxide (DMSO), absorption spectra of AK3-1 exhibits vibrational structure with the relative intensity of 0-0 sub-band being higher than that for the 0-1 sub-band. In an aqueous phase the dye absorption band undergoes hypsochromic shift relative to DMSO due to H-aggregation of the dye. The interaction of AK3-1 with the native and fibrillar insulin was followed by the decrease of monomer band and the enhancement of H-dimer band. To evaluate the relative contributions of the monomeric and aggregated forms, the absorption spectra of the protein-bound dye were deconvoluted using the asymmetric log-normal (LN) function. The analysis of the set of fitting parameters provides evidence for the protein-induced AK3-1 self-association into the head-to-head dimers, with the magnitude of this effect being much more pronounced for fibrillar protein form. The molecular docking studies showed that the AK3-1 monomer tends to associate with the specific arrangement of side chains in the ß-sheet formed by L17 leucine residues (of the insulin B-chain), located on the dry steric zipper interface of the fibril, while the dye dimers form stable complexes with the amyloid groove formed by the residues Q15 and E17 of the A-chain, and located on the wet interface of the fibril. The latter binding site is more easily accessible and is additionally stabilized by the electrostatic interactions between the positively charged dye and the E17 residue. This binding mode seems to be prevailing over that for the AK3-1 monomers. Based on the results obtained, AK3-1 may be recommended as a prospective amyloid marker complementary to the classical amyloid reporters Thioflavin T and Congo Red.