Strongylocentrotus purpuratus spindle tubulin. I. Characteristics of its polymerization and depolymerization in vitro.

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Abstract
Tubulin was extracted from spindles isolated from embryos of the sea urchin S. prupuratus, repolymerized in vitro, and purified through 3 cycles of temperature-dependent assembly and disassembly. In addition to the tubulin, these preparations contain a protein of 80 kdaltons and a small but a variable amount of actin. At 37.degree. C, the tubulin polymerizes with a critical concentration of 0.15-0.2 mg/ml into smooth-walled polymers which contain predominantly 14 protofilaments. Removal of the 80 kdalton protein and the actin by DEAE-chromatography does not change the critical concentration for polymerization. At 15.degree. C, which is within the range of physiological temperatures for S. purpuratus embryos, the spindle tubulin will self-assemble, but the rate of total polymer formation is very slow, requiring hours in the test tube. This rate can be increased by shearing the polymerizing microtubules, creating more ends for assembly, indicating that the slow rate of polymer formation is due to a slow rate of self-initiation. If spindle tubulin is polymerized at 37.degree. C and then lowered to 15.degree. C, some polymer will be retained, the percentage of which depends on the protein concentration. Spindle tubulin from S. purpuratus will assemble at 37.degree. C with a low critical concentration for polymerization in the absence of detectable microtubule-associated proteins and will self-assemble and maintain steady state levels of polymer at physiological temperatures.