Stabilization of hepatic colchicine-binding activity by organic acids

Previous work has shown that the total hepatic tubulin pool and the hepatic microtubule-derived tubulin pool do not have identical [³H]colchicine binding properties. Rapid loss of colchicine-binding activity was noted in the microtubule-derived fractions of liver tubulin. Furthermore, quantitative determination of the total and polymerized tubulin in the liver by the [³H]colchicine-binding assay was hampered by rapid and unequal loss of binding sites under assay conditions. The organic acids, glutamate and glucose 1-phosphate, have been shown to stabilize calf brain tubulin against loss of colchicine-binding sites. Therefore, these compounds were tested as possible protecting agents against loss of colchicine binding activity of liver tubulin. It was found that these agents stabilized liver tubulin under [³H]colchicine-binding conditions. Additional experiments showed that these agents also prevented the rapid loss of colchicine-binding activity that occurred when purified brain tubulin was exposed to liver supernates. These results suggest that the inclusion of the organic acids, glutamate and glucose 1-phosphate, may modify the time decay properties of liver tubulin in solution. Further, these data suggest that these protecting agents may be of analytical value in [³H]colchicine-binding assay systems for liver tubulin.