Zabala J C, Fontalba A, Avila J
Departamento de Biologia Molecular, Facultad de Medicina, Universidad de Cantabria, Spain.
J Cell Sci. 1996 Jun;109 ( Pt 6):1471-8. doi: 10.1242/jcs.109.6.1471.
Tubulins contain a glycine-rich loop, that has been implicated in microtubule dynamics by means of an intramolecular interaction with the carboxy-terminal region. As a further extension of the analysis of the role of the carboxy-terminal region in tubulin folding we have mutated the glycine-rich loop of tubulin subunits. An alpha-tubulin point mutant with a T150-->G substitution (the corresponding residue present in beta-tubulin) was able to incorporate into dimers and microtubules. On the other hand, four beta-tubulin point mutants, including the G148-->T substitution, did not incorporate into dimers, did not release monomers, but were able to form C900 and C300 complexes (intermediates in the process of tubulin folding). Three other mutants within this region (which approximately encompasses residues 137-152) were incapable of forming dimers and C300 complexes but gave rise to the formation of C900 complexes. These results suggest that tubulin goes through two sequential folding states during the folding process, first in association with TCP1-complexes (C900) prior to the transfer to C300 complexes. It is this second step that implies binding/hydrolysis of GTP, reinforcing our previous proposed model for tubulin folding and assembly.
微管蛋白含有一个富含甘氨酸的环，该环通过与羧基末端区域的分子内相互作用参与微管动力学。作为对羧基末端区域在微管蛋白折叠中作用分析的进一步扩展，我们对微管蛋白亚基的富含甘氨酸的环进行了突变。一个具有T150→G替代（β-微管蛋白中存在的相应残基）的α-微管蛋白点突变体能够组装成二聚体和微管。另一方面，四个β-微管蛋白点突变体，包括G148→T替代，不能组装成二聚体，不能释放单体，但能够形成C900和C300复合物（微管蛋白折叠过程中的中间体）。该区域内的其他三个突变体（大致涵盖残基137 - 152）不能形成二聚体和C300复合物，但能导致C900复合物的形成。这些结果表明，微管蛋白在折叠过程中经历两个连续的折叠状态，首先与TCP1复合物（C900）结合，然后再转移到C300复合物。正是这第二步意味着GTP的结合/水解，加强了我们先前提出的微管蛋白折叠和组装模型。
