Developmental and Biochemical Analysis of Chick Brain Tubulin Heterogeneity

Abstract: Tubulin, isolated from brain tissue of chicks at different stages during late embryonic and early posthatched development by ion‐exchange chromatography and by in vitro microtubule reassembly, was analyzed by high‐resolution isoelectric focusing and by two‐dimensional polyacrylamide gel electrophoresis. Similar results were obtained with tubulins purified by the two methods. Sixteen isoelectric species of tubulin that differ in apparent net charge under denaturing conditions were detected by isoelectric focusing. By two‐dimensional polyacrylamide gel electrophoresis, the chick brain tubulins were resolved into at least seven forms of α and 10 forms of β tubulin. The number and relative proportions of the maltiple brain tubulins were modulated during development. Since there are only four α tubulin and four β tubulin genes in chickens, posttranslational modification of the tubulins must play a prominent role in the heterogeneity. Analysis of isotubulin distributions through cycles of microtubule assembly and disassembly indicated that the tubulins differ very little, if at all, in their capacity to assemble into microtubules. Therefore, the chemical differences that distinguish the multiple tubulins have very little structural impact on the protein surface areas involved in microtubule formation. Partial fractionation of the multiple tubulins during ion‐exchange chromatography was observed, suggesting that it may be possible to isolate individual native tubulin variants for biochemical studies.