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Drug discovery and design based on molecular mimicry

基于分子模拟的药物发现和设计

基本信息

批准号：
11557190
负责人：
NAKAMURA Yoshikazu
金额：
$ 8.58万
依托单位：
The University of Tokyo
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
1999
资助国家：
日本
起止时间：
1999 至 2001
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11557190/
关键词：
RNA SELEX in vitro evolution aptamer molecular mimicry antagonist 癌抑制翻訳開始因子インターロイキン

项目摘要

The recent progress in molecular and structural biology of translational apparatus has uncovered a novel biological phenomenon of macromolecular, mimicry between protein and tRNA. The 3D structures of translational elongation factor EF- G and ribosome recycling factor RRF are shown to mimic a tRNA shape. Another, even more sophisticated, example may be polypeptide release factors that are shown to possess a tripeptide anticodon' to decipher stop codons in mRNA, showing an impressive mimic of tRNA function. Clearly, this concept should provide us with a novel clue to design or selection of therapeutic RNA molecules that mimic protein targets.The systematic evolution of ligands by exponential enrichment (SELEX) method is based on the in vitro selection oligo-nucleotide; ligands from large random-sequence libraries by repeated reactions of DNA transcription, RNA selection and RT-PCR amplification. The selected oligo-nucleotide ligands are called 'aptamer', which has. High affinity and specificity to the target molecule. We have initiated SELEX experiments using mammalian translation initiation factors including eIF4G and eIF4A. Several RNA aptamers that acquired the strong and specific binding capacity for these proteins were successfully raised. One class of eIF4A aptamers exhibits a high affinity with.el4A1 and severely inhibits the ATPasc activity, suggesting that they probably dock the catalytic pocket. Furthermore, these aptamers inhibit can dependent protein synthesis in a cell free translation system. This inhibition was reversed by addition of purified eIF4A1 suggesting that RNA aptamers to eIF4A1 inhibits its activity in the cell lysate.Importantly the abnormality in the protein level or the activity of either initiation factor is known cause cell proliferation. Hence, from the therapeutic point of view, it is of great interest whether or not these RNA aptamers can suppress these malignant cells.

翻译装置的分子和结构生物学的最新进展揭示了一种新的生物学现象，即蛋白质和tRNA之间的大分子拟态。翻译延伸因子EF-G和核糖体回收因子RRF的三维结构模拟tRNA形状。另一个更复杂的例子可能是多肽释放因子，它被证明具有三肽反密码子，可以破译mRNA中的终止密码子，显示出令人印象深刻的对tRNA功能的模拟。显然，这个概念应该为我们设计或选择模拟蛋白质靶标的治疗性RNA分子提供新的线索。指数富集法(SELEX)的系统进化基于体外选择寡核苷酸；通过DNA转录、RNA选择和RT-PCR扩增的重复反应从大的随机序列文库中选择配体。所选择的寡核苷酸配体被称为‘适配子’，它具有。对靶分子具有较高的亲和力和特异性。我们已经使用包括eIF4G和eIF4A在内的哺乳动物翻译启动因子启动了SELEX实验。成功地培育了几个与这些蛋白质具有较强的特异性结合能力的RNA适配子。一类eIF4A适配子与el4A1显示出高亲和力，并严重抑制ATPasc活性，表明它们可能对接催化口袋。此外，这些适配子抑制细胞自由翻译系统中依赖于CaN的蛋白质合成。这种抑制作用可被纯化的eIF4A1逆转，这表明eIF4A1的RNA适配子抑制其在细胞裂解液中的活性。重要的是，蛋白质水平的异常或任一启动因子的活性已知会导致细胞增殖。因此，从治疗的角度来看，这些RNA适配子能否抑制这些恶性细胞是人们非常感兴趣的。