Real-Time Observation of the Interaction between Thioflavin T and an Amyloid Protein by Using High-Sensitivity Rheo-NMR

Naoto Iwakawa; Daichi Morimoto; Erik Walinda; Yasushi Kawata; Masahiro Shirakawa; Kenji Sugase

doi:10.3390/ijms18112271

Real-Time Observation of the Interaction between Thioflavin T and an Amyloid Protein by Using High-Sensitivity Rheo-NMR

Int J Mol Sci. 2017 Oct 28;18(11):2271. doi: 10.3390/ijms18112271.

Authors

Naoto Iwakawa¹, Daichi Morimoto², Erik Walinda³, Yasushi Kawata⁴, Masahiro Shirakawa⁵, Kenji Sugase⁶

Affiliations

¹ Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Kyoto-Daigaku Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. iwakawa.naoto.46w@st.kyoto-u.ac.jp.
² Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Kyoto-Daigaku Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. morimoto@moleng.kyoto-u.ac.jp.
³ Department of Molecular and Cellular Physiology, Graduate School of Medicine, Kyoto University, Yoshida Konoe-cho, Sakyo-ku, Kyoto 606-8501, Japan. walinda.erik.6e@kyoto-u.ac.jp.
⁴ Department of Chemistry and Biotechnology, Graduate School of Engineering, Tottori University, 4-101 Koyama-cho Minami, Tottori 680-8552, Japan. kawata@bio.tottori-u.ac.jp.
⁵ Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Kyoto-Daigaku Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. shirakawa@moleng.kyoto-u.ac.jp.
⁶ Department of Molecular Engineering, Graduate School of Engineering, Kyoto University, Kyoto-Daigaku Katsura, Nishikyo-ku, Kyoto 615-8510, Japan. sugase@moleng.kyoto-u.ac.jp.

Abstract

Amyloid fibril formation is associated with numerous neurodegenerative diseases. To elucidate the mechanism of fibril formation, the thioflavin T (ThT) fluorescence assay is widely used. ThT is a fluorescent dye that selectively binds to amyloid fibrils and exhibits fluorescence enhancement, which enables quantitative analysis of the fibril formation process. However, the detailed binding mechanism has remained unclear. Here we acquire real-time profiles of fibril formation of superoxide dismutase 1 (SOD1) using high-sensitivity Rheo-NMR spectroscopy and detect weak and strong interactions between ThT and SOD1 fibrils in a time-dependent manner. Real-time information on the interaction between ThT and fibrils will contribute to the understanding of the binding mechanism of ThT to fibrils. In addition, our method provides an alternative way to analyze fibril formation.

Keywords: Rheo-NMR; SOD1; amyloid fibrils; molecular interactions; real-time observation; thioflavin T.

MeSH terms

Amyloid / metabolism
Amyloid / ultrastructure
Amyloidogenic Proteins / chemistry
Amyloidogenic Proteins / metabolism*
Amyloidogenic Proteins / ultrastructure
Benzothiazoles
Magnetic Resonance Spectroscopy*
Protein Binding
Superoxide Dismutase-1 / metabolism
Superoxide Dismutase-1 / ultrastructure
Thiazoles / chemistry
Thiazoles / metabolism*

Substances

Amyloid
Amyloidogenic Proteins
Benzothiazoles
Thiazoles
thioflavin T
Superoxide Dismutase-1