Rapid and Simple Ribozymic Aminoacylation Using Three Conserved Nucleotides
Rapid and Simple Ribozymic Aminoacylation Using Three Conserved Nucleotides
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DOI：
10.1021/ja809419f
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发表时间：
2009-04-15
影响因子：
15
通讯作者：
Yarus, M.
中科院分区：
文献类型：
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作者：
Chumachenko, N. V.;Novikov, Y.;Yarus, M.
选择扩增在随机化的RNA中发现新的RNA酶（核酶），其侧翼序列不变（引物互补序列）。在反应前精确去除X-引物，在三个循环中从PheAMP中选择氨酰化，仅使用三个保守核苷酸产生活性RNA（k（cat）= 12-20 min（-1）），独立于二价离子起作用。这个异常简单的RNA活性位点鼓励了通过基于分子力学的自由能最小化来研究反应。在此基础上，我们提出了一种RNA催化转氨酰化的化学途径。位点模型还预测了新的特征，L-立体选择性，2 '-区域选择性，氨基酸侧链的独立性，和磷酸化的活化基团，随后得到验证。同样的选择也表明，腺苷酸的RNA氨酰化比CoA硫酯简单，可能合理化腺苷酸的翻译激活。这种活性位点的简单性表明了小核酶的一般途径。
Selection-amplification finds new RNA enzymes (ribozymes) among randomized RNAs with flanking unvaried sequences (primer complements). Precise removal of X-primer before reaction selected aminoacylation from PheAMP in three cycles, yielding active RNAs (k(cat) = 12-20 min(-1)) using only three conserved nucleotides, acting independently of divalent ions. This unusually simple RNA active site encouraged study of the reaction via molecular mechanics-based free energy minimization. On this basis, we suggest a chemical path for RNA-catalyzed transaminoacylation. Site modeling also predicted new features, L-stereoselectivity, 2'-regioselectivity, independence of amino acid side chain, and phosphorylated activating group, that were subsequently verified. The same selection also showed that RNA aminoacylation from adenylate is simpler than from CoA thioester, potentially rationalizing translational activation by adenylates. The simplicity of this active site suggests a general route to small ribozymes.