MONOVALENT CATION INDUCED STRUCTURAL TRANSITIONS IN TELOMERIC DNAS - G-DNA FOLDING INTERMEDIATES
MONOVALENT CATION INDUCED STRUCTURAL TRANSITIONS IN TELOMERIC DNAS - G-DNA FOLDING INTERMEDIATES
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DOI：
10.1021/bi00232a013
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发表时间：
1991-05-07
期刊：
影响因子：
2.9
通讯作者：
PROSSER, JK
中科院分区：
文献类型：
--
作者：
HARDIN, CC;HENDERSON, E;PROSSER, JK
端粒DNA由富含G和C的链组成，它们总是被极化，使得富含G的链延伸超过双链体的3'端以形成12-16个碱基的突出端。 这些悬垂链可以在体外自缔合形成分子内结构，这些结构具有几种不寻常的物理性质和至少一个共同特征，即存在非沃森-克里克G.G碱基对。 术语“G-DNA”是为这类结构创造的（Cech，1988）。 凝胶电泳、亚氨基质子核磁共振和圆二色性（CD）结果表明，将抗衡离子从钠离子改变为钾离子，（在20 mM磷酸盐缓冲液中）特异性诱导来自四膜虫的富含G的端粒DNA中的构象转变，d（T2 G4）4（TET 4），这导致从分子内物质到明显的多链结构的变化，伴随着碱基对的熔融温度增加> 25度，如通过亚氨基质子NMR信号的损失所监测的。 NMR半选择性自旋晶格弛豫速率测量和HPLC尺寸排阻色谱研究表明，在20 mM磷酸钾（pH 7）缓冲液（KP）中，TET 4的长度约为在20 mM磷酸钠（pH 7）缓冲液（NaP）中获得的形式的两倍，Na+和K+的混合物产生两种形式的混合物，其种群取决于阳离子的比例。 由于已知K+和NH 4+稳定poly[r（I）4]的平行链四链体结构，我们推断多链结构是四链体。 我们的研究结果表明，离子相互作用的特定差异可以导致端粒DNA分子内发夹样或四链体的物种和分子间四链体结构之间的开关，所有这些都涉及G.G碱基配对相互作用。 我们提出了一个模型，其中双链体或发夹形式的G-DNA折叠中间体形成的1-，2-或4-链四链体结构。 在该模型中，一价阳离子通过两种不同的机制稳定双链体和四链体形式，即与堆叠平面鸟嘌呤“四重体”碱基组装体中的羰基的抗衡缩合和八面体配位。 取代四膜虫序列的每个重复序列中的一个鸟苷残基以产生人类端粒DNA d（T2 AG 3）4，导致K+依赖性稳定化的效果较低。 因此，通过改变序列来减弱离子依赖性稳定。 在添加沃森-克里克（WC）互补链后，仅Na+稳定的结构快速解离以形成WC双螺旋。 这表明在某些情况下，K+稳定的G-DNA结构可以在动力学上优于WC DNA。
Telomeric DNA consists of G- and C-rich strands that are always polarized such that the G-rich strand extends past the 3' end of the duplex to form a 12-16-base overhang. These overhanging strands can self-associate in vitro to form intramolecular structures that have several unusual physical properties and at least one common feature, the presence of non-Watson-Crick G.G base pairs. The term "G-DNA" was coined for this class of structures (Cech, 1988). On the basis of gel electrophoresis, imino proton NMR, and circular dichroism (CD) results, we find that changing the counterions from sodium to potassium (in 20 mM phosphate buffers) specifically induces conformational transitions in the G-rich telomeric DNA from Tetrahymena, d(T2G4)4 (TET4), which results in a change from the intramolecular species to an apparent multistranded structure, accompanied by an increase in the melting temperature of the base pairs of > 25-degrees, as monitored by loss of the imino proton NMR signals. NMR semiselective spin-lattice relaxation rate measurements and HPLC size-exclusion chromatography studies show that in 20 mM potassium phosphate (pH 7) buffer (KP) TET4 is approximately twice the length of the form obtained in 20 mM sodium phosphate (pH 7) buffer (NaP) and that mixtures of Na+ and K+ produce mixtures of the two forms whose populations depend on the ratio of the cations. Since K+ and NH4+ are known to stabilize a parallel-stranded quadruplex structure of poly[r(I)4], we infer that the multistranded structure is a quadruplex. Our results indicate that specific differences in ionic interactions can result in a switch in telomeric DNAs between intramolecular hairpin-like or quadruplex-containing species and intermolecular quadruplex structures, all of which involve G.G base pairing interactions. We propose a model in which duplex or hairpin forms of G-DNA are folding intermediates in the formation of either 1-, 2-, or 4-stranded quadruplex structures. In this model monovalent cations stabilize the duplex and quadruplex forms via two distinct mechanisms, counterion condensation and octahedral coordination to the carbonyl groups in stacked planar guanine "quartet" base assemblies. Substituting one of the guanosine residues in each of the repeats of the Tetrahymena sequence to give the human telomeric DNA, d(T2AG3)4, results in less effective K+-dependent stabilization. Thus, the ion-dependent stabilization is attenuated by altering the sequence. Upon addition of the Watson-Crick (WC) complementary strand, only the Na+-stabilized structure dissociates quickly to form a WC double helix. This demonstrates that under some circumstances the K+-stabilized G-DNA structure can be kinetically preferred over WC DNA.