The reconstitution of microtubules from purified calf brain tubulin

James Lee, Serge N. Timasheff

Research output: Contribution to journalArticle

162 Citations (Scopus)

Abstract

The in vitro reconstitution of calf brain tubulin, purified by the method of Weisenberg et al. [(1968), Biochemistry 7, 4466-4479; (1970), Biochemistry 9, 4110-4116] as modified by Lee et al. [(1973), J. Biol. Chem. 248, 7253-7262], was successful in a medium consisting of 10-2 M sodium phosphate, 10-4 M GTP, and concentrations of magnesium ions ranging from 0.5 to 16 × 10-3 M at 37°. Filaments resembling native microtubules were formed. The filaments are in equilibrium with the associating species of tubulin and the equilibrium can be shifted to depolymerization by lowering the temperature to 20°. Fila-ment formation is inhibited by calcium ions which also cause disassembly of the formed filaments. The effects of calcium ion can be reversed by the addition of [ethylenebis(oxyethylenenitrilo)]tetraacetic acid. The formation of filaments is favored by the presence of 3.4 M glycerol; only twisted abnormal filaments are observed in the presence of 1 M sucrose. The high molecular weight components observed in the sodium dodecyl sulfate polyacrylamide gel electrophoresis patterns of many tubulin preparations were shown not to be essential for the formation of the filaments.

Original languageEnglish (US)
Pages (from-to)5183-5187
Number of pages5
JournalBiochemistry
Volume14
Issue number23
StatePublished - 1975
Externally publishedYes

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Tubulin
Microtubules
Brain
Biochemistry
Ions
Calcium
Depolymerization
Electrophoresis
Sodium Dodecyl Sulfate
Glycerol
Sucrose
Polyacrylamide Gel Electrophoresis
Molecular Weight
Molecular weight
Temperature
Acids

ASJC Scopus subject areas

  • Biochemistry

Cite this

Lee, J., & Timasheff, S. N. (1975). The reconstitution of microtubules from purified calf brain tubulin. Biochemistry, 14(23), 5183-5187.

The reconstitution of microtubules from purified calf brain tubulin. / Lee, James; Timasheff, Serge N.

In: Biochemistry, Vol. 14, No. 23, 1975, p. 5183-5187.

Research output: Contribution to journalArticle

Lee, J & Timasheff, SN 1975, 'The reconstitution of microtubules from purified calf brain tubulin', Biochemistry, vol. 14, no. 23, pp. 5183-5187.
Lee, James ; Timasheff, Serge N. / The reconstitution of microtubules from purified calf brain tubulin. In: Biochemistry. 1975 ; Vol. 14, No. 23. pp. 5183-5187.
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