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Protein Expr Purif. 2003 Nov;32(1):83-8.

Purification of brain tubulin through two cycles of polymerization-depolymerization in a high-molarity buffer.

Author information

1
European Molecular Biology Laboratory, Cell Biology and Biophysics Programme, Meyerhofstrasse 1, 69117 Heidelberg, Germany.

Abstract

Microtubules can be assembled in vitro from purified alpha/beta tubulin heterodimers in the presence of GTP. Tubulin is routinely obtained from animal brain tissue through repetitive cycles of polymerization-depolymerization, followed by ion-exchange chromatography to remove any contaminating microtubule-associated proteins and motors. Here, we show that only two cycles of polymerization-depolymerization of pig brain tubulin in the presence of a high-molarity PIPES buffer allow the efficient removal of contaminating proteins and production of a high-concentration tubulin solution. The proposed protocol is rapid and yields more active tubulin than the traditional ion-exchange chromatography-based procedures.

PMID:
14680943
DOI:
10.1016/S1046-5928(03)00218-3
[Indexed for MEDLINE]

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