Journal
PROTEIN EXPRESSION AND PURIFICATION
Volume 32, Issue 1, Pages 83-88Publisher
ACADEMIC PRESS INC ELSEVIER SCIENCE
DOI: 10.1016/S1046-5928(03)00218-3
Keywords
microtubule; tubulin; tubulin polymerization; brain tubulin; microtubule-associated protein
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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. (C) 2003 Elsevier Inc. All rights reserved.
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