Novobiocin and related coumarins and depletion of heat shock protein 90-dependent signaling proteins
Novobiocin and related coumarins and depletion of heat shock protein 90-dependent signaling proteins
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DOI：
10.1093/jnci/92.3.242
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发表时间：
2000-02-02
影响因子：
10.3
通讯作者：
Neckers, L
中科院分区：
文献类型：
--
作者：
Marcu, MG;Schulte, TW;Neckers, L
背景：热休克蛋白 90 (Hsp90) 与多种致癌蛋白激酶（例如 p185(erbB2)、p60(v-src) 和 Raf-1）相互作用并使其稳定，并且是突变 p53 蛋白的稳定性和显性失活功能所必需的。两种不相关的抗生素，格尔德霉素和根赤霉素，特异性结合 Hsp90 的非典型核苷酸结合袋，该位点与细菌 DNA 旋转酶 B 的三磷酸腺苷 (ATP) 结合域具有同源性。这种相互作用导致与 Hsp90 相互作用的蛋白质不稳定。由于旋转酶 B 的核苷酸结合位点是香豆素抗生素（例如新生霉素）的靶标，因此我们研究了这些药物是否也可以与 Hsp90 相互作用并影响其活性。方法：我们使用固定化新生霉素、格尔德霉素或根赤霉素从细胞裂解物中分离内源性 Hsp90 或体外翻译的 Hsp90 缺失片段。在体外和体内评估了香豆素类抗生素新生霉素、氯生霉素和香豆霉素 A1 对与 Hsp90 相互作用的几种蛋白质的影响。结果：Hsp90 与固定化新生霉素的结合受到可溶性香豆素和 ATP 的竞争，但不受到格尔德霉素或根赤霉素的竞争。羧基末端Hsp90片段结合固定化新生霉素但不结合固定化格尔德霉素，而结合格尔德霉素的氨基末端片段不结合新生霉素。所有三种香豆素均显着降低 p185(erbB2)、p60(v-src)、Raf-1 和突变 p53 的细胞水平。此外，新生霉素降低了接受该药物治疗的小鼠脾脏中的 Raf-1 水平。结论：这些香豆素抗生素，特别是新生霉素，是其他耐受性较差的 Hsp90 靶向药物的第一代替代品。新生霉素与 Hsp90 的独特相互作用确定了该蛋白质上易于受到小分子药理干扰的额外位点。
Background: Heat shock protein 90 (Hsp90) interacts with and stabilizes several oncogenic protein kinases (e.g., p185(erbB2), p60(v-src), and Raf-1) and is required for the stability and dominant-negative function of mutated p53 protein. Two unrelated antibiotics, geldanamycin and radicicol, bind specifically to an atypical nucleotide-binding pocket of Hsp90, a site that shares homology with the adenosine triphosphate (ATP)-binding domain of bacterial DNA gyrase B. This interaction leads to destabilization of proteins that interact with Hsp90. Since the nucleotide-binding site of gyrase B is targeted by coumarin antibiotics (e.g., novobiocin), we investigated whether these drugs can also interact with Hsp90 and affect its activity. Methods: We used immobilized novobiocin, geldanamycin, or radicicol to isolate either endogenous Hsp90 from cell lysates or Hsp90 deletion fragments translated in vitro. Effects of the coumarin antibiotics novobiocin, chlorobiocin, and coumermycin A1 on several proteins interacting with Hsp90 were assessed in vitro and in vivo. Results: Hsp90 binding to immobilized novobiocin was competed by soluble coumarins and ATP but not by geldanamycin or radicicol. A carboxy-terminal Hsp90 fragment bound immobilized novobiocin but not immobilized geldanamycin, while a geldanamycin-binding amino-terminal fragment did not bind novobiocin. All three coumarins markedly reduced cellular levels of p185(erbB2), p60(v-src), Raf-l, and mutated p53. Furthermore, novobiocin reduced Raf-l levels in the spleens of mice treated with the drug. Conclusions: These coumarin antibiotics, particularly novobiocin, represent a first-generation alternative to other Hsp90-targeting drugs that are not as well tolerated. Novobiocin's unique interaction with Hsp90 identifies an additional site on this protein amenable to pharmacologic interference with small molecules.