Abstract
Publisher Summary This chapter presents the methods for isolating microtubule protein from hog and beef brain, along with the procedures for fractionating microtubule protein into separate tubulin and microtubule-associated protein (MAP) fractions. Microtubules are prominent components of the cytoplasmic matrix and perform important functions as cytoskeletal elements for the determination of cell shape and as key elements in intracellular motility such as mitosis and the translocation of cell organelles. The purification methods described in the chapter are based on the temperature dependence of microtubule polymerization. Microtubule assembly is an equilibrium process involving free tubulin subunits and microtubule polymers. Warm temperature (37°C) shifts the equilibrium to favor polymer, whereas cold temperature (0°C) shifts the equilibrium to favor free dissociated subunits. An assembly–disassembly procedure that does not employ glycerol was originally used to purify microtubules from extracts of hog brain. MAPs bind to tubulin in large quantity and co-purify in constant stoichiometry to tubulin through several successive cycles of in vitro assembly–disassembly.
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