Studies of interaction between cyanine dye T-284 and fibrillar alpha-synuclein.
A key feature of Parkinson's disease is the formation and accumulation of amyloid fibrils of the natively unfolded protein α-synuclein (ASN) inside neurons. Recently we have proposed novel sensitive monomethinecyanine dye T-284 as fluorescent probe for quantitative detection of ASN amyloid fibrils. In this study the T-284 dye complex with ASN fibril was characterized by means of fluorescence anisotropy, atomic force microscopy and time-resolved fluorescence techniques to give further insights into the mode of dye interaction with amyloid fibrils. The fluorescence anisotropy of T-284 was shown to noticeably increase upon addition of aggregated proteins indicating on stable dye/amyloid fibril complex formation. AFM imaging of fibrillar wild-type ASN revealed differences in heights between ASN fibrils alone and in presence of the T-284 dye (6.37 ± 1.0 nm and 8.0 ± 1.1 nm respectively), that is believed to be caused by embedding of T-284 dye molecules in the "binding channel" running along the fibril. Fluorescence decay analysis of the T-284 in complexes with fibrillar ASN variants revealed the fluorescence lifetime values for T-284/fibril complexes to be an order of magnitude higher as compared to the free dye. Also, the fluorescence decay of free T-284 was bi-exponential, while dye bound to protein yields tri-exponential decay. We suppose that in complexes with fibrillar ASN variants T-284 dye might exist in different "populations" due to interaction with fibrils in different conformers and ways. The exact binding mode of T-284 with ASN fibrils needs further studies. Studied parameters of dye/amyloid fibril complexes are important for the characterization and screening of newly-developed amyloid-sensitive dyes.
Volkova
Kateryna D
KD
Department of Combinatorial Chemistry, Institute of Molecular Biology and Genetics of the National Academy of Sciences of Ukraine, 150 Zabolotnogo St., 03143, Kyiv, Ukraine.
Kovalska
Vladyslava B
VB
Yu Losytskyy
Mykhaylo
M
Veldhuis
Gertjan
G
Segers-Nolten
G M J
GM
Tolmachev
Olexiy I
OI
Subramaniam
Vinod
V
Yarmoluk
Sergiy M
SM
D002232:Carbocyanines / Q000737:chemistry*; D005454:Fluorescence Polarization; D005456:Fluorescent Dyes / Q000737:chemistry*; D018625:Microscopy, Atomic Force; D015394:Molecular Structure; D051844:alpha-Synuclein / Q000737:chemistry*
D002232:Carbocyanines; D005456:Fluorescent Dyes; C556877:T-284 dye; D051844:alpha-Synuclein