Bode Claudia J, Gupta Mohan L, Reiff Emily A, Suprenant Kathy A, Georg Gunda I, Himes Richard H
Department of Molecular Biosciences and the Department of Medicinal Chemistry, University of Kansas, Lawrence, Kansas 66045, USA.
Biochemistry. 2002 Mar 26;41(12):3870-4. doi: 10.1021/bi0121611.
Paclitaxel (Taxol) and the epothilones are antimitotic agents that promote the assembly of mammalian tubulin and stabilization of microtubules. The epothilones competitively inhibit the binding of paclitaxel to mammalian brain tubulin, suggesting that the two types of compounds share a common binding site in tubulin, despite the lack of structural similarities. It is known that paclitaxel does not stabilize microtubules formed in vitro from Saccharomyces cerevisiae tubulin; thus, it would be expected that the epothilones would not affect yeast microtubules. However, we found that epothilone A and B do stimulate the formation of microtubules from purified yeast tubulin. In addition, epothilone B severely dampens the dynamics of yeast microtubules in vitro in a manner similar to the effect of paclitaxel on mammalian microtubules. We used current models describing paclitaxel and epothilone binding to mammalian beta-tubulin to explain why paclitaxel apparently fails to bind to yeast tubulin. We propose that three amino acid substitutions in the N-terminal region and at position 227 in yeast beta-tubulin weaken the interaction of the 3'-benzamido group of paclitaxel with the protein. These results also indicate that mutagenesis of yeast tubulin could help define the sites of interaction with paclitaxel and the epothilones.
紫杉醇（泰素）和埃坡霉素是抗有丝分裂剂，可促进哺乳动物微管蛋白组装并稳定微管。埃坡霉素竞争性抑制紫杉醇与哺乳动物脑微管蛋白的结合，这表明尽管这两类化合物缺乏结构相似性，但它们在微管蛋白中共享一个共同的结合位点。已知紫杉醇不能稳定由酿酒酵母微管蛋白在体外形成的微管；因此，可以预期埃坡霉素不会影响酵母微管。然而，我们发现埃坡霉素A和B确实能刺激从纯化的酵母微管蛋白形成微管。此外，埃坡霉素B在体外严重抑制酵母微管的动态变化，其方式类似于紫杉醇对哺乳动物微管的作用。我们使用描述紫杉醇和埃坡霉素与哺乳动物β-微管蛋白结合的当前模型来解释为什么紫杉醇显然无法与酵母微管蛋白结合。我们提出，酵母β-微管蛋白N端区域和第227位的三个氨基酸取代削弱了紫杉醇3'-苯甲酰胺基团与该蛋白的相互作用。这些结果还表明，酵母微管蛋白的诱变有助于确定与紫杉醇和埃坡霉素的相互作用位点。
