Human telomeric quadruplex conformations studied by pulsed EPR.
Human telomeric quadruplex conformations studied by pulsed EPR.
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DOI：
10.1002/anie.200902146
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发表时间：
2009-12
影响因子：
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通讯作者：
Vijay R. Singh;M. Azarkh;T. Exner;J. Hartig;M. Drescher
中科院分区：
文献类型：
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作者：
Vijay R. Singh;M. Azarkh;T. Exner;J. Hartig;M. Drescher
富含鸟苷的核酸折叠成称为四链体的四链结构。11.21 染色体末端的端粒重复引起了人们的极大兴趣。在人类中，它们由六聚体 GGGTIA 重复序列组成，其 3' 末端由 100 至 200 个核苷酸的单链突出端组成。！'1 由于其在折叠和未折叠状态之间切换的潜力，四链体结构的形成被怀疑在端粒维持和细胞周期控制中发挥重要作用。14.51 与双链体结构相反，四链体在拓扑特征方面表现出高度的多态性，例如单条链的方向和环的连接性。例如，已知人类端粒重复序列会根据参数（例如由四链体协调的单价离子类型和核苷酸序列的轻微变化）采用截然不同的构象。16 81 NMR 光谱和晶体学研究已经提供了令人印象深刻的人类端粒四链体的高分辨率结构。生物物理学研究表明这些结构具有相似的稳定性，并提供了越来越多的证据证明其中一些折叠在生理条件下共存。不幸的是，高分辨率方法无法在接近生理条件下破译这些结构的确切性质，因为这些技术需要单一物种的存在。诸如圆二色性 (CD) 光谱之类的生物物理方法提供的信息非常丰富，但结果往往被过度解释191，并且最近产生了有争议的结果，如下所述。在此，我们利用双硝基氧修饰的端粒序列结合双频脉冲电子顺磁共振（EPR）方法研究了近生理条件下的四链体构象（图1a）。双电子电子共振（DEER 或 PELDOR）光谱非常适合区分不同的构象，因为它允许距离
Guanosine rich nucleic acids fold into four stranded struc tures called quadruplexes.11.21 The telomeric repeats at the end of the chromosomes, have generated much interest. In humans, they are composed of the hexameric GGGTIA repeat with the 3' ends consisting of a 100 to 200 nucleotide single stranded overhang.!'1 As a result of their potential to switch between folded and unfolded state, the formation of quadruplex structures is suspected to play important roles in telomere maintenance and cell cycle control.14.51 In contrast to duplex structures, quadruplexes show a high degree of polymorphism with respect to topological features, such as the orientation of individual strands and the connectivity of the loops. For example, the human telomeric repeat is known to adopt drastically different conformations depending on parameters such as the type of monovalent ions coordinated by the quadruplex and the slight changes in the nucleotide seq uence.16 81 NMR spectroscopic and crystallographic studies have delivered impressive high resolution structures of human telomeric quadruplexes. Biophysical studies indicate similar stabilities of these structures and provide increasing evidence of the co existence of some of these folds under physiological conditions. Unfortunately, the high resolution methods are not able to decipher the exact nature of these structures under near physi.ological conditions since these techniques require the presence of single species. Biophysical methods such as circular dichroism (CD) spectroscopy are very informative but the results tend to be over interpretedl91 and have produced controversial results recently as discussed below. Herein, we investigate quadruplex conformations under near physiological conditions utilizing double nitroxide modified telomeric sequences in combination with a two frequency pulsed electron paramagnetic resonance (EPR) method (Figure 1 a). Double electron electron resonance (DEER or PELDOR) spectroscopy is ideally suited to distinguish between the different conformations since it allows distance