Structure-Function Analysis of Yeast Tubulin

摘要: Microtubules play essential roles in a wide variety of cellular processes including cell division, motility, and vesicular transport. Microtubule function depends on the polymerization dynamics tubulin specific interactions between diverse microtubule-associated proteins. To date, investigation structural functional properties mutants has been limited by inability to obtain protein from overexpression systems, heterogeneous mixture isotypes typically isolated higher eukaryotes. The budding yeast, Saccharomyces cerevisiae, emerged as leading system for structure-function analysis. Yeast cells encode single beta-tubulin gene can be engineered express just one two alpha isotypes. Moreover, yeast allows site-directed modification genes at endogenous loci expressed under native promoter regulatory elements. These advantageous features provide homogeneous controlled environment analysis consequences mutations. Here, we present techniques generate site-specific mutations diploid haploid cells, assess ability mutated support viability, measure overall microtubule stability, define changes parameters dynamic instability. We also outline strategies determine whether disrupt with Microtubule-based functions are well defined, which observed related role microtubules processes.