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Arch Biochem Biophys. 2018 Aug 1;651:13-20. doi: 10.1016/j.abb.2018.05.019. Epub 2018 May 24.

Dissection of the deep-blue autofluorescence changes accompanying amyloid fibrillation.

Author information

1
International Laser Center, M.V. Lomonosov Moscow State University, Leninskie Gory, Moscow, 119991, Russia.
2
Faculty of Physics, M.V. Lomonosov Moscow State University, Leninskie Gory, Moscow, 119991, Russia.
3
Skolkovo Institute of Science and Technology, 143025 Skolkovo, Moscow region, Russia.
4
A.N. Bach Institute of Biochemistry, Federal Research Center "Fundamentals of Biotechnology" of the Russian Academy of Sciences, Moscow, 119071, Russia; Faculty of Biology, M.V. Lomonosov Moscow State University, Leninskie Gory, Moscow, 119991, Russia.
5
National Research Center "Kurchatov Institute", 1 pl. Akademika Kurchatova, Moscow, 123182, Russia.
6
Skolkovo Institute of Science and Technology, 143025 Skolkovo, Moscow region, Russia; Institute for Energy Problems of Chemical Physics Russian Academy of Sciences Leninskij pr. 38 k.2, 119334, Moscow, Russia.
7
Department of Molecular Medicine and USF Health Byrd Alzheimer's Research Institute, Morsani College of Medicine, University of South Florida, Tampa, Florida, 33612, USA; Institute for Biological Instrumentation of the Russian Academy of Sciences, Institutskaya str., 7, Pushchino, Moscow Region, 142290, Russia.
8
Faculty of Physics, M.V. Lomonosov Moscow State University, Leninskie Gory, Moscow, 119991, Russia. Electronic address: shirshin@lid.phys.msu.ru.

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.

KEYWORDS:

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

PMID:
29803394
DOI:
10.1016/j.abb.2018.05.019

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