Demonstration of three dopamine molecules bound to &amp;lt;i&amp;gt;α&amp;lt;/i&amp;gt;-Synuclein: Implication of oligomerization at the initial stage

Sakurako Shimotakahara,Sayaka Kado,Kenji Uéda,Yuuki Shiroyama,Mai Akai,Yoichi Shibusawa,Takashi Fujimoto,Mitsuru Tashiro,Takaya Onoue,Hiroko Seki,Tomoya Machinami

doi:10.4236/jbpc.2012.32017

Abstract

α-Synuclein is the major component of the filamentous Lewy bodies and Lewy neurites that define neuropathological features and dementia with Lewy bodies. To investigate the role of dopamine (DA) in α-synuclein fibrillation, the structural propensities to form oligomers at the initial stage fibrillation were studied using size exclusion chromatography and various biophysical techniques. Interactions with DA were observed for wild-type α-synuclein and its mutants, A30P, E46K and A53T, using electrospray ionization mass spectrometry (ESI-MS). The results of ESI-MS indicate that an intact α-synuclein, which was not oxidized, had an ability to bind with three molecules of DA at the initial stage. Furthermore, upon binding to DA, α-synuclein oligomerizes to higher molecular weight species. These oligomers are structurally different from amyloid fibrils, as confirmed by thioflavin T and CD analysis.

Highlights

Α-Synuclein is the major component of the filamentous Lewy bodies and Lewy neurites that define neuropathological features and dementia with Lewy bodies
In order to observe the effects of DA in amyloid fibril formation of α-synucleins, an accumulation of amyloid was monitored by fluorescence spectroscopy using a ThT binding assay
These results indicate that DA significantly inhibited the amyloid fibril formation of the wild-type α-synuclein and its three mutants

Summary

Introduction

Α-Synuclein is the major component of the filamentous Lewy bodies and Lewy neurites that define neuropathological features and dementia with Lewy bodies. To investigate the role of dopamine (DA) in α-synuclein fibrillation, the structural propensities to form oligomers at the initial stage fibrillation were studied using size exclusion chromatography and various biophysical techniques. Upon binding to DA, α-synuclein oligomerizes to higher molecular weight species These oligomers are structurally different from amyloid fibrils, as confirmed by thioflavin T and CD analysis. In order to investigate the detailed oligomer association mechanism of α-synuclein at the initial stage, interactions between α-synuclein and DA have been studied using various biochemical and biophysical techniques. The measurements using an electrospray ionization mass spectrometry (ESI-MS), size exclusion chromatography, fluorescence and CD have been carried out for the wild-type α-synuclein and three mutants, A30P, E46K and A53T, to evaluate the effect of these mutations in oligomerization.

Methods

Results

Conclusion