Yu, Hongtao2015-01-142015-01-142012-122012-12-04December 2https://hdl.handle.net/2152.5/ETD-UTSWMED-2012-12-60https://hdl.handle.net/2152.5/1474Stu2p/XMAP215/Dis1 family proteins are evolutionarily conserved regulatory factors that use alpha/beta-tubulin-interacting TOG (tumor overexpressed gene) domains to catalyze fast microtubule growth. Catalysis requires that these polymerases discriminate between unpolymerized and polymerized forms of alpha/beta-tubulin, but how they do so has remained unclear. In this study, we first introduce the polymerization blocked mutants of alpha/beta-tubulins that we developed as unique tools for biochemical studies of alpha/beta-tubulins to avoid the difficulties that has arisen from the self-assembly tendency of tubulins, then we report the structure of the TOG1 domain from Stu2p bound to the plus end polymerization blocked yeast alpha/beta-tubulin we created to facilitate crystallization. Our structure and further biochemical characterizations of the TOG1:alpha/beta-tubulin complex showed that TOG1 binds alpha/beta-tubulin in a way that excludes equivalent binding of a second TOG domain. Furthermore, TOG1 preferentially binds a “curved” conformation of alpha/beta-tubulin that cannot be incorporated into microtubules, contacting α- and β-tubulin surfaces that do not participate in microtubule assembly. Conformation-selective interactions with alpha/beta-tubulin explain how TOG-containing polymerases discriminate between unpolymerized and polymerized forms of alpha/beta-tubulin, and how they selectively recognize the growing end of the microtubule.application/pdfenMicrotubule-Associated ProteinsMicrotubulesSaccharomyces cerevisiae ProteinsTubulinThesis2015-01-14900281614