Structure-function relationships in yeast tubulins
Structure-function relationships in yeast tubulins
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DOI：
10.1091/mbc.11.5.1887
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发表时间：
2000-05-01
影响因子：
3.3
通讯作者：
Botstein, D
中科院分区：
文献类型：
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作者：
Richards, KL;Anders, KR;Botstein, D
产生了一套全面的簇状丙氨酸突变，系统地改变酿酒酵母的主要 α 微管蛋白基因 TUB1。观察到多种表型，包括对微管不稳定药物苯菌灵的超敏感性和耐药性、致死性以及对冷和温度敏感的致死性。许多对苯菌灵最敏感的 tube1 等位基因与 tube3 Delta 结合具有综合致死性，这支持了苯菌灵超敏感性是微管不稳定性和/或 cr-微管蛋白量不足的粗略衡量标准的观点。将系统性的 tube1 突变与先前描述的类似的 tube2 突变组一起放置到基于牛微管蛋白三维结构的酵母 α-β-微管蛋白二聚体模型上。该模型揭示了 β-微管蛋白核心中苯菌灵结合的潜在位点。引起冷敏感性的突变残基集中在相邻亚基之间的横向和纵向界面处。影响微管结合蛋白 Bim1p 结合的残基在模型中的 α-微管蛋白的外表面形成一个大斑块。最后，突变的位置表明，靠近 α-β 界面可能是发现五个可行的 tube1 等位基因与苯菌灵抗性但完全可行的 tube2-201 等位基因的合成致死性的原因。
A comprehensive set of clustered charged-to-alanine mutations was generated that systematically alter TUB1, the major alpha-tubulin gene of Saccharomyces cerevisiae. A variety of phenotypes were observed, including supersensitivity and resistance to the microtubule-destabilizing drug benomyl, lethality, and cold- and temperature-sensitive lethality. Many of the most benomyl-sensitive tub1 alleles were synthetically lethal in combination with tub3 Delta, supporting the idea that benomyl supersensitivity is a rough measure of microtubule instability and/or insufficiency in the amount of cr-tubulin. The systematic tub1 mutations were placed, along with the comparable set of tub2 mutations previously described, onto a model of the yeast alpha-beta-tubulin dimer based on the three-dimensional structure of bovine tubulin. The modeling revealed a potential site for binding of benomyl in the core of beta-tubulin. Residues whose mutation causes cold sensitivity were concentrated at the lateral and longitudinal interfaces between adjacent subunits. Residues that affect binding of the microtubule-binding protein Bim1p form a large patch across the exterior-facing surface of alpha-tubulin in the model. Finally, the positions of the mutations suggest that proximity to the alpha-beta interface may account for the finding of synthetic lethality of five viable tub1 alleles with the benomyl-resistant but otherwise entirely viable tub2-201 allele.