Detection and quantification of protein-microtubules interactions using green fluorescent protein photoconversion.
We present an in vitro system to analyze quantitatively the interactions of green fluorescent protein (GFP)-tagged recombinant proteins with microtubules. This method relies on photoconversion of GFP and time-lapse microscopy. Specific interactions can be detected and binding kinetics can be determined rapidly and accurately. This method provides an alternative to classical in vitro microtubule-binding assays to analyze microtubule-associated proteins binding to microtubules. It has the potential to be extended to study interactions of proteins or multi-protein complexes with different biopolymers like actin microfilaments or organelle membranes.
Brunet
Stéphane
S
Biologie du Développement, CNRS UMR 7622 bat C-5ème, 9, Quai St Bernard, 75252 Paris Cedex 05, France. sbrunet@snv.jussieu.fr
Zimmermann
Timo
T
Reynaud
Emmanuel G
EG
Vernos
Isabelle
I
Karsenti
Eric
E
Pepperkok
Rainer
R
D049452:Green Fluorescent Proteins; D006801:Humans; D008869:Microtubule-Associated Proteins / Q000378:metabolism*; D008870:Microtubules / Q000378:metabolism*; D013194:Staining and Labeling
D008869:Microtubule-Associated Proteins; D049452:Green Fluorescent Proteins