A distance ruler for RNA using EPR and site-directed spin labeling
A distance ruler for RNA using EPR and site-directed spin labeling
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DOI：
10.1016/j.chembiol.2004.04.013
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发表时间：
2004-07-01
影响因子：
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通讯作者：
DeRose, VJ
中科院分区：
文献类型：
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作者：
Kim, NK;Murali, A;DeRose, VJ
作为使用连续波（CW）EPR光谱测量RNA分子距离的基本模型研究，研究了定点自旋标记的10-mer RNA双链体和具有各种自旋间距离的HIV-1 TAR RNA基序。将自旋标记物连接到双链体中适当放置的尿苷的2 '-NH 2位置，并通过分子动力学模拟（MD）和傅立叶去卷积方法（FD）测量自旋间距离[13]。基于MD，10-mer双链体具有10埃至30埃范围内的自旋间距离;然而，仅对于预测的10-21埃的自旋间距离的RNA观察到CW EPR谱的偶极线加宽，而对于超过25埃的距离未观察到。通过测量凸起区域中两个核苷酸之间的距离来监测TAR（反式激活响应区）RNA在不同二价金属离子存在和不存在下的构象变化。从FD方法和MD计算得到的预测的自旋间距离匹配以及模型RNA双链体和TAR RNA预测的结构变化。这些结果表明，使用EPR光谱的距离测量是一个潜在的强大的方法，以帮助预测RNA分子的结构。
As a basic model study for measuring distances in RNA molecules using continuous wave (CW) EPR spectroscopy, site-directed spin-labeled 10-mer RNA duplexes and HIV-1 TAR RNA motifs with various interspin distances were examined. The spin labels were attached to the 2'-NH2 positions of appropriately placed uridines in the duplexes, and interspin distances were measured from both molecular dynamics simulations (MD) and Fourier deconvolution methods (FD) [13]. The 10-mer duplexes have interspin distances ranging from 10 Angstrom to 30 Angstrom based on MD; however, dipolar line broadening of the CW EPR spectrum is only observed for the RNAs for predicted interspin distances of 10-21 Angstrom and not for distances over 25 Angstrom. The conformational changes in TAR (transactivating responsive region) RNA in the presence and in the absence of different divalent metal ions were monitored by measuring distances between two nucleotides in the bulge region. The predicted interspin distances obtained from the FD method and those from MD calculations match well for both the model RNA duplexes and the structural changes predicted for TAR RNA. These results demonstrate that distance measurement using EPR spectroscopy is a potentially powerful method to help predict the structures of RNA molecules.