3'-(p-azidobenzamido)taxol photolabels the N-terminal 31 amino acids of beta-tubulin.
3'-(p-azidobenzamido)taxol photolabels the N-terminal 31 amino acids of beta-tubulin.
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DOI：
10.1016/s0021-9258(17)41836-9
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发表时间：
1994-02
期刊：
影响因子：
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通讯作者：
SRINIVASA K. Rao;N. Krauss;J. M. Heerding;C. Swindell;I. Ringel;G. Orr;S. Horwitz
中科院分区：
文献类型：
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作者：
SRINIVASA K. Rao;N. Krauss;J. M. Heerding;C. Swindell;I. Ringel;G. Orr;S. Horwitz
紫杉醇具有不寻常的化学结构、独特的作用机制，并在人类恶性肿瘤中表现出活性。它是唯一在微管聚合物上具有结合位点的抗肿瘤剂。紫杉醇与微管聚合物的相互作用导致形成抗解聚的稳定的细胞微管束。虽然微管，特别是β-微管蛋白，是紫杉醇的靶点已经变得很明显，但没有关于药物结合位点的信息。在这份报告中，我们证明了3 '-（p-叠氮苯甲酰氨基）紫杉醇，一种类似物与紫杉醇的生物活性，共价结合的微管-药物复合物照射后的β-微管蛋白的N-末端结构域。紫杉醇与[3 H] 3 '-（对叠氮基苯甲酰氨基）-紫杉醇竞争结合，表明光亲和类似物和紫杉醇在相同或重叠位点结合。甲酸裂解[3 H] 3 '-（p-叠氮基苯甲酰氨基）-紫杉醇光标记的β-微管蛋白和随后的蛋白质序列和质量分析已经鉴定N-末端31个氨基酸为[3 H] 3'-（p-叠氮基苯甲酰氨基）紫杉醇光掺入的主要位点。
Taxol possesses an unusual chemical structure, a unique mechanism of action, and demonstrated activity in human malignancies. It is the only antitumor agent that has a binding site on the microtubule polymer. The interaction of Taxol with the microtubule polymer results in the formation of stable bundles of cellular microtubules that are resistant to depolymerization. Although it has become evident that the microtubule, specifically beta-tubulin, is the target for Taxol, no information is available on the binding site for the drug. In this report, we demonstrate that 3'-(p-azidobenzamido)taxol, an analogue with similar biological activities as Taxol, covalently binds to the N-terminal domain of beta-tubulin after irradiation of the microtubule-drug complex. Taxol competes with [3H]3'-(p-azidobenzamido)-taxol binding, suggesting that the photoaffinity analog and Taxol are binding at the same or overlapping sites. Formic acid cleavage of [3H]3'-(p-azidobenzamido)-taxol-photolabeled beta-tubulin and subsequent protein sequence and mass analyses have identified the N-terminal 31 amino acids as the major site for [3H]3'-(p-azidobenzamido)taxol photoincorporation.