In vitro assembly of pure tubulin into microtubules in the absence of microtubule-associated proteins and glycerol.

Abstract

Microtubule protein from porcine cerebrum was fractionated into pure tubulin and microtubule-associated proteins by chromatography on phosphocellulose. In agreement with previous studies, pure tubulin did not form microtubules to a significant extent at 37.degree. C in normal assembly buffers, characterized by a low concentration of Mg2+ ions. If the Mg2+ concentration was raised to approximately 10 mM, rapid and extensive self-assembly of pure tubulin into microtubules was observed, provided the tubulin concentration was above 2.5 mg/ml. At a protein concentration of 3 mg/ml, the lag period was 1.5 min and the assembly process was virtually complete after 6 min at 37.degree. C. These microtubules were like normal microtubules, sensitive to Ca2+ colchicine and low temperature.