Drug design based on the complex structure of HIV-Tat and its RNA aptamer

基于HIV-Tat及其RNA适体复杂结构的药物设计

基本信息

批准号：
12470487
负责人：
KATAHIRA Masato
金额：
$ 5.38万
依托单位：
Yokohama National University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2002
项目状态：
已结题

项目摘要

An RNA aptamer containing two binding sites exhibits extremely high affinity to the HIV Tat protein. We have now determined the structure of the aptamer complexed with argininamide, the simplest analogue of the Tat protein, by NMR. We found that two argininamide molecules bind per aptamer. Two adjacent U : A : U base triples were formed, which widens the major groove to make space for the two argininamide molecules. The argininamide molecules bound to the G bases through hydrogen bonds. The binding is stabilized through stacking interactions.The structure of the aptamer complexed with a Tat-derived argmine-rich peptide was also characterized. The Tat-derived peptide bound to the aptamer in a 1: 1 molar ratio. In spite of the difference in stoichiometry, similarity was noted between the argininamide- and peptide-bound forms of the aptamer in chemical shift perturbations of the aptamer upon complex formation and intermolecular contacts. These results suggest that two different arginine residues of the peptide interact with the two binding sites of the aptamer in the same way as two argininamide molecules do. Simultaneous interactions of the aptamer with two arginine residues of Tat could explain its high affinity compared to the authentic TAR RNA. The formation of the two adjacent base triples makes the simultaneous interactions possible by creating space for the accommodation of two arginine residues and linking residues of Tat, and also contributes to the stabilization of the binding through stacking interactions.The elucidated structure of the aptamer gives clues to rationally design a new aptamer that exhibits even higher affinity to Tat. The way to design a new aptamer with a less side effect is also provided by the elucidated structure.

含有两个结合位点的RNA适体对HIV Tat蛋白表现出极高的亲和力。我们现在已经通过 NMR 确定了与 Tat 蛋白最简单的类似物精氨酰胺复合的适体的结构。我们发现每个适体结合两个精氨酰胺分子。形成两个相邻的 U : A : U 碱基三元组，这加宽了主沟，为两个精氨酰胺分子腾出了空间。精氨酰胺分子通过氢键与 G 碱基结合。通过堆积相互作用使结合稳定。还表征了与 Tat 衍生的富含精胺的肽复合的适体的结构。 Tat 衍生肽以 1:1 摩尔比与适体结合。尽管化学计量存在差异，但在复合物形成和分子间接触时适体的化学位移扰动方面，适体的精氨酰胺结合形式和肽结合形式之间存在相似性。这些结果表明，肽的两个不同的精氨酸残基与适体的两个结合位点的相互作用方式与两个精氨酰胺分子的相互作用相同。适体与 Tat 的两个精氨酸残基同时相互作用可以解释其与真实 TAR RNA 相比的高亲和力。两个相邻碱基三元组的形成通过为两个精氨酸残基和连接 Tat 残基创造空间，使得同时相互作用成为可能，并且还有助于通过堆叠相互作用稳定结合。适配体的阐明结构提供了线索合理设计新的适体，对Tat表现出更高的亲和力。阐明的结构还提供了设计副作用较小的新适体的方法。