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Method Article
Didier Portran
Nachury lab
Corresponding Author
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.
Keywords
Tubulin, protofilaments, FRET, fluorimeter, longitudinal interactions, lateral interactions
The authors declare no competing financial interests.
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Biochemistry
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)
Method Article
Didier Portran
Nachury lab
Corresponding Author
Version 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.
Loading...
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)