Direct Detection of α-Synuclein Dimerization Dynamics: Single-Molecule Fluorescence Analysis. Academic Article uri icon

Overview

MeSH

  • Point Mutation

MeSH Major

  • Protein Multimerization
  • alpha-Synuclein

abstract

  • The aggregation of α-synuclein (α-Syn) is linked to Parkinson's disease. The mechanism of early aggregation steps and the effect of pathogenic single-point mutations remain elusive. We report here a single-molecule fluorescence study of α-Syn dimerization and the effect of mutations. Specific interactions between tethered fluorophore-free α-Syn monomers on a substrate and fluorophore-labeled monomers diffusing freely in solution were observed using total internal reflection fluorescence microscopy. The results showed that wild-type (WT) α-Syn dimers adopt two types of dimers. The lifetimes of type 1 and type 2 dimers were determined to be 197 ± 3 ms and 3334 ± 145 ms, respectively. All three of the mutations used, A30P, E46K, and A53T, increased the lifetime of type 1 dimer and enhanced the relative population of type 2 dimer, with type 1 dimer constituting the major fraction. The kinetic stability of type 1 dimers (expressed in terms of lifetime) followed the order A30P (693 ± 14 ms) > E46K (292 ± 5 ms) > A53T (226 ± 6 ms) > WT (197 ± 3 ms). Type 2 dimers, which are more stable, had lifetimes in the range of several seconds. The strongest effect, observed for the A30P mutant, resulted in a lifetime 3.5 times higher than observed for the WT type 1 dimer. This mutation also doubled the relative fraction of type 2 dimer. These data show that single-point mutations promote dimerization, and they suggest that the structural heterogeneity of α-Syn dimers could lead to different aggregation pathways. Copyright © 2015 Biophysical Society. Published by Elsevier Inc. All rights reserved.

publication date

  • April 21, 2015

has subject area

  • Point Mutation
  • Protein Multimerization
  • alpha-Synuclein

Research

keywords

  • Journal Article

Identity

Language

  • eng

PubMed Central ID

  • PMC4407253

Digital Object Identifier (DOI)

  • 10.1016/j.bpj.2015.03.010

PubMed ID

  • 25902443

Additional Document Info

start page

  • 2038

end page

  • 2047

volume

  • 108

number

  • 8