Probing amylin fibrillation at an early stage via a tetracysteine-recognising fluorophore

Shih Ting Wang; Yiyang Lin; Chia Chen Hsu; Nadav Amdursky; Christopher D. Spicer; Molly M. Stevens

doi:10.1016/j.talanta.2017.05.015

Probing amylin fibrillation at an early stage via a tetracysteine-recognising fluorophore

Shih Ting Wang, Yiyang Lin, Chia Chen Hsu, Nadav Amdursky, Christopher D. Spicer, Molly M. Stevens^*

^*Corresponding author for this work

Materials Science and Engineering

Research output: Contribution to journal › Article › peer-review

14 Scopus citations

Abstract

Amyloid fibrillation is a nucleation-dependent process known be involved in the development of more than 20 progressive and chronic diseases. The detection of amyloid formation at the nucleation stage can greatly advance early diagnoses and treatment of diseases. In this work, we developed a new assay for the early detection of amylin fibrillation using the biarsenical dye 4,5-bis(1,3,2-dithiarsolan-2-yl)fluorescein (FlAsH), which could recognise tetracysteine motifs and transform from non-fluorescent form into strongly fluorescent complexes. Due to the close proximity of two cysteine residues within the hydrophilic domain of amylin, a non-contiguous tetracysteine motif can form upon amylin dimerisation or oligomerisation, which can be recognised by FlAsH and emit strong fluorescence. This enables us to report the nucleation-growth process of amylin without modification of the protein sequence. We showed that the use of this assay not only allowed the tracking of initial nucleation events, but also enabled imaging of amyloid fibrils and investigation of the effects of amyloid inhibitor/modulator toward amylin fibrillation.

Original language	English (US)
Pages (from-to)	44-50
Number of pages	7
Journal	Talanta
Volume	173
DOIs	https://doi.org/10.1016/j.talanta.2017.05.015
State	Published - Oct 1 2017

Funding

S.-T.W. and C.-C.H thank the support of the Top University Strategic Alliance Ph.D. scholarship from Taiwan. N.A. was supported by Marie Curie actions FP7 through the Intra-European Marie Curie Fellowship \u201CConPilus\u201D under grant agreement no. 623123. C.D.S. and M.M.S. acknowledge the Engineering and Physical Sciences Research Council (EPSRC) grant \u201CBio-functionalised nanomaterials for ultrasensitive biosensing\u201D (EP/K020641/1). M.M.S. also acknowledges support from the ERC Seventh Framework Programme Consolidator grant \u201CNaturale CG\u201D under Grant Agreement No. 616417 and the EPSRC Interdisciplinary Research Centre (IRC) \u201CEarly-Warning Sensing Systems for Infectious Diseases\u201D (EP/K031953/1). Raw data is available online at DOI: 10.5281/zenodo.259524.

Keywords

Amylin
Amyloid inhibitor
Early detection
Fibrillation
FlAsH
Glycosaminoglycan

ASJC Scopus subject areas

Analytical Chemistry

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10.1016/j.talanta.2017.05.015

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title = "Probing amylin fibrillation at an early stage via a tetracysteine-recognising fluorophore",

abstract = "Amyloid fibrillation is a nucleation-dependent process known be involved in the development of more than 20 progressive and chronic diseases. The detection of amyloid formation at the nucleation stage can greatly advance early diagnoses and treatment of diseases. In this work, we developed a new assay for the early detection of amylin fibrillation using the biarsenical dye 4,5-bis(1,3,2-dithiarsolan-2-yl)fluorescein (FlAsH), which could recognise tetracysteine motifs and transform from non-fluorescent form into strongly fluorescent complexes. Due to the close proximity of two cysteine residues within the hydrophilic domain of amylin, a non-contiguous tetracysteine motif can form upon amylin dimerisation or oligomerisation, which can be recognised by FlAsH and emit strong fluorescence. This enables us to report the nucleation-growth process of amylin without modification of the protein sequence. We showed that the use of this assay not only allowed the tracking of initial nucleation events, but also enabled imaging of amyloid fibrils and investigation of the effects of amyloid inhibitor/modulator toward amylin fibrillation.",

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N2 - Amyloid fibrillation is a nucleation-dependent process known be involved in the development of more than 20 progressive and chronic diseases. The detection of amyloid formation at the nucleation stage can greatly advance early diagnoses and treatment of diseases. In this work, we developed a new assay for the early detection of amylin fibrillation using the biarsenical dye 4,5-bis(1,3,2-dithiarsolan-2-yl)fluorescein (FlAsH), which could recognise tetracysteine motifs and transform from non-fluorescent form into strongly fluorescent complexes. Due to the close proximity of two cysteine residues within the hydrophilic domain of amylin, a non-contiguous tetracysteine motif can form upon amylin dimerisation or oligomerisation, which can be recognised by FlAsH and emit strong fluorescence. This enables us to report the nucleation-growth process of amylin without modification of the protein sequence. We showed that the use of this assay not only allowed the tracking of initial nucleation events, but also enabled imaging of amyloid fibrils and investigation of the effects of amyloid inhibitor/modulator toward amylin fibrillation.

AB - Amyloid fibrillation is a nucleation-dependent process known be involved in the development of more than 20 progressive and chronic diseases. The detection of amyloid formation at the nucleation stage can greatly advance early diagnoses and treatment of diseases. In this work, we developed a new assay for the early detection of amylin fibrillation using the biarsenical dye 4,5-bis(1,3,2-dithiarsolan-2-yl)fluorescein (FlAsH), which could recognise tetracysteine motifs and transform from non-fluorescent form into strongly fluorescent complexes. Due to the close proximity of two cysteine residues within the hydrophilic domain of amylin, a non-contiguous tetracysteine motif can form upon amylin dimerisation or oligomerisation, which can be recognised by FlAsH and emit strong fluorescence. This enables us to report the nucleation-growth process of amylin without modification of the protein sequence. We showed that the use of this assay not only allowed the tracking of initial nucleation events, but also enabled imaging of amyloid fibrils and investigation of the effects of amyloid inhibitor/modulator toward amylin fibrillation.

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Probing amylin fibrillation at an early stage via a tetracysteine-recognising fluorophore

Abstract

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Keywords

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