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Method Article
Measurement of tubulin oligomers self-assembly by FRET.
Didier Portran
Nachury lab
Corresponding Author
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This work is licensed under a CC BY-NC 3.0 License. Read Full License
Microtubules nucleation, dynamic instability and mechanical properties are regulated by tubulin-tubulin longitudinal and lateral interactions. It is thus crucial to develop assays that can investigate potential modifications of these interactions due to tubulin post-translational modifications or mutations. Here we describe a detailed protocol for a method to monitor tubulin oligomerization at a step before MT nucleation occurs, using a FRET-based assay. Briefly, the principle of the technique is to measure over time the fluorescence emitted by a FRET pair of fluorophores when tubulin dimers interact with each other below the critical concentration for microtubules self-assembly. This technique can also be used to monitor more specifically lateral interactions between protofilaments each labelled with one FRET pair of fluorophores. The experiment can be completed within 2-3 hours.
Keywords
Tubulin, protofilaments, FRET, fluorimeter, longitudinal interactions, lateral interactions
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Associated Publications
Tubulin acetylation protects long-lived microtubules against mechanical ageing
by Didier Portran, Laura Schaedel, Zhenjie Xu, +2
Nature Cell Biology (09 March, 2017)
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This is a preprint. It has not completed peer review.
Method Article
Measurement of tubulin oligomers self-assembly by FRET.
Didier Portran
Nachury lab
Corresponding Author
Badges
Prescreen
This manuscript passed our Prescreen assessment
Complete author information
Appropriate declaration statements
Potential risks to human health
Prescreen
History
CURRENT STATUS: Posted
Version 1
Posted 09 Mar, 2017
No community comments so far
Metrics
Comments: 0
HTML Views: ...
Subject Areas
Biochemistry
License:
This work is licensed under a CC BY-NC 3.0 License. Read Full License
Microtubules nucleation, dynamic instability and mechanical properties are regulated by tubulin-tubulin longitudinal and lateral interactions. It is thus crucial to develop assays that can investigate potential modifications of these interactions due to tubulin post-translational modifications or mutations. Here we describe a detailed protocol for a method to monitor tubulin oligomerization at a step before MT nucleation occurs, using a FRET-based assay. Briefly, the principle of the technique is to measure over time the fluorescence emitted by a FRET pair of fluorophores when tubulin dimers interact with each other below the critical concentration for microtubules self-assembly. This technique can also be used to monitor more specifically lateral interactions between protofilaments each labelled with one FRET pair of fluorophores. The experiment can be completed within 2-3 hours.
Associated Publications
Tubulin acetylation protects long-lived microtubules against mechanical ageing
by Didier Portran, Laura Schaedel, Zhenjie Xu, +2
Nature Cell Biology (09 March, 2017)