Dissection of the deep-blue autofluorescence changes accompanying amyloid fibrillation

Tatiana N. Tikhonova, Nataliya R. Rovnyagina, Alexander Ya Zherebker, Nikolai N. Sluchanko, Anna A. Rubekina, Anton S. Orekhov, Eugene N. Nikolaev, Victor V. Fadeev, Vladimir N. Uversky, Evgeny A. Shirshin

    Research output: Contribution to journalArticlepeer-review

    31 Citations (Scopus)

    Abstract

    Pathogenesis of numerous diseases is associated with the formation of amyloid fibrils. Extrinsic fluorescent dyes, including Thioflavin T (ThT), are used to follow the fibrillation kinetics. It has recently been reported that the so-called deep-blue autofluorescence (dbAF) is changing during the aggregation process. However, the origin of dbAF and the reasons for its change remain debatable. Here, the kinetics of fibril formation in model proteins were comprehensively analyzed using fluorescence lifetime and intensity of ThT, intrinsic fluorescence of proteinaceous fluorophores, and dbAF. For all systems, intensity enhancement of the dbAF band with similar spectral parameters (∼350 nm excitation; ∼450 nm emission) was observed. Although the time course of ThT lifetime (indicative of protofibrils formation) coincided with that of tyrosine residues in insulin, and the kinetic changes in the ThT fluorescence intensity (reflecting formation of mature fibrils) coincided with changes in ThT absorption spectrum, the dbAF band started to increase from the beginning of the incubation process without a lag-phase. Our mass-spectrometry data and model experiments suggested that dbAF could be at least partially related to oxidation of amino acids. This study scrutinizes the dbAF features in the context of the existing hypotheses about the origin of this spectral band.

    Original languageEnglish
    Pages (from-to)13-20
    Number of pages8
    JournalArchives of Biochemistry and Biophysics
    Volume651
    DOIs
    Publication statusPublished - 1 Aug 2018

    Keywords

    • Deep-blue autofluorescence
    • Fibrillation kinetics
    • Fibrils
    • Intrinsic fluorescence
    • Protein oxidation
    • Thioflavin T

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