Hairpin ribozymes with four-way helical junctions mediate intracellular RNA ligation.
Hairpin ribozymes with four-way helical junctions mediate intracellular RNA ligation.
复制标题
具有四向螺旋连接的发夹核酶介导细胞内 RNA 连接。
DOI：
10.1006/jmbi.2001.4713
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发表时间：
2001
期刊：
影响因子：
--
通讯作者：
Fedor,MJ
中科院分区：
文献类型：
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作者：
Yadava,RS;Choi,AJ;Lebruska,LL;Fedor,MJ
几乎所有的RNA介导的反应都需要在不同的RNA构象之间进行转换。生物反应的复杂性会掩盖体内特定的构象变化，细胞内环境的重要特征很难在体外复制。然而，简单的RNA自切和连接反应提供了一个独特的机会，可以直接在活细胞中测量特定RNA构象转变的动力学和平衡。发夹状核酶结合了天然的四向螺旋连接，在体内迅速自我切割，但只有在切割产物迅速解离时才能自我切割。当切割产物保持结合在稳定的碱基配对螺旋中时，切割速率下降，这提供了结合产物经历重新连接的证据。这些结果首次提供了细胞内核酶反应的详细动力学描述，包括切割、连接和产物解离速率。在体内测量的动力学和平衡参数与在接近细胞内离子环境的条件下体外测得的相同反应的值很好地吻合，但不是完全一致。
Virtually all RNA-mediated reactions require transitions among alternative RNA conformations. The complexity of biological reactions can obscure specific conformational changes in vivo and important features of the intracellular environment are difficult to reproduce in vitro. However, simple RNA self-cleavage and ligation reactions offer a unique opportunity to measure the kinetics and equilibria of specific RNA conformational transitions directly in living cells. Hairpin ribozymes that incorporate the natural four-way helical junction self-cleave rapidly in vivo, but only when cleavage products dissociate rapidly. Cleavage rates fall when cleavage products remain bound in stable base-paired helices, providing evidence that bound products undergo re-ligation. These results provide the first detailed kinetic description of an intracellular ribozyme reaction that includes cleavage, ligation and product dissociation rates. Kinetic and equilibrium parameters measured in vivo correspond well, but not perfectly, with values measured for the same reactions in vitro under conditions that approximate an intracellular ionic environment.