Human brain tubulin purification: decrease in soluble tubulin with age
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The soluble tubulin of human cerebral cortex, as assessed by [3H]colchicine binding of the 100,000 g supernatant fraction, decreases drastically with age, 75 percent from age 0 to age 90. There is also a considerably lower concentration of high molecular weight proteins in the soluble fraction of postmortem human cerebral cortex than in that of nonhuman species. Human brain tubulin can be polymerized into microtubules with DEAE-dextran. The DEAE-dextran induced microtubules are stable to cold temperature (4 degrees) and calcium. However, in the presence of 1 M glutamate, the microtubules become cold labile and depolymerize at 4 degrees. Thus we have developed a novel method for purifying polymerization competent tubulin from fresh or frozen human cerebral cortex. Human brain tubulin purified by our novel method is very similar to tubulin from the brains of other mammals in molecular weight, amino acid composition, polymerization-depolymerization parameters, and structural dimensions of the microtubules formed.
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