The Dickerson-Drew B-DNA dodecamer revisited at atomic resolution
The Dickerson-Drew B-DNA dodecamer revisited at atomic resolution
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DOI：
10.1021/ja9832919
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发表时间：
1999-01-20
影响因子：
15
通讯作者：
Egli, M
中科院分区：
文献类型：
--
作者：
Tereshko, V;Minasov, G;Egli, M
我们对 B 型 DNA 的了解大部分源于对寡脱氧核苷酸 CGCGAATTCGCG（即所谓的 Dickerson-Drew dodecamer (DDD)）的结构研究。它的晶体结构提供了第一张右手 DNA 双螺旋的详细图像。 1 基于该结构和相关十二聚体解决的问题包括碱基序列和结构的相互依赖性、2 主链柔性、3 溶剂化、4 弯曲性和弯曲性、3、5 药物结合、6 以及包装力 7 和结晶条件 8 对 DNA 结构的影响。 DDD 双链体的有趣特征是 AATT 区域中小沟的狭窄以及该沟中水分子的脊柱。然而，X 射线晶体学迄今为止未能揭示抗衡离子（特别是一价和二价金属阳离子）对 B-DNA 结构的影响。原因之一似乎是 DDD 晶体结构的分辨率有限（平均约 2.3 Å）。根据分子动力学 (MD) 模拟，有人认为 Na+ 离子可以侵入小沟中的电负性“AT 口袋”，并以部分占有率驻留在那里。 A-tract DNA 的 9 NMR 溶液实验为小沟中存在 Mn2+ 离子提供了证据。 10 g1。天然 DDD11 的 5 Å 结构和含有化学修饰胸苷 12 的 12mer 促使我们进行了最先进的晶体学实验，目的是最大限度地提高 DDD 结构的分辨率并了解有关双链体离子环境的更多信息。在这里，我们以 1.1 Å 的分辨率报告了 DDD 晶体结构的详细信息，这是迄今为止 B-DNA 双链体获得的最高分辨率。导致分辨率显着提高的因素包括过去几年寡核苷酸合成和纯化方面的改进。 13 然而，在这方面，原始结晶条件的修改、14 晶体的适当冷冻以及第三代同步加速器源的数据收集15 可能更为重要。数据收集和细化17统计数据总结于表1中。每个不对称单元存在三种有序的Mg2+离子，两种六水合物（Mg1和Mg3）和一种五水合物络合物（Mg2）（图1；a、b等指定对称配合物）。 Mg1 位于主沟中，靠近双相体的一端。 11, 12 离子通过配位水接触相对链的残基 G2 和 G22 的 N7 和 O6 边缘。它还桥接相邻分子的 P6c 和 P7c 的 O2P 氧，并稳定 P2 和 P7c (6.73 Å) 之间紧密的双链体间接触。这种离子相互作用可能导致 DDD 双链体不对称地扭结到主凹槽中（图 2）。 8 P20 和 P12d (6.68 Å) 以及 P10 和 P18d (6.24 Å) 之间存在额外的紧密横向双链体间接触。 Mg2 直接与磷酸盐 P19 的 O1P 配位，此外还通过其水配体之一与磷酸盐 P12d 的 O1P 形成 H 键。同样，Mg3 通过相同的配位水分别桥接磷酸盐 P10 和 P18d 的氧 O2P 和 O1P（图 1）。 Mg2 可稳定分子两端磷酸盐 P12 和 P24 (5.59 Å) 之间的紧密接触。因此，Mg2+ 离子位于双链体之间的端到端重叠附近，这是 DDD 晶格的一个特殊特征。 7 如图 1 和 2 所示，Mg2 和 Mg3 还可缓解磷酸盐 P10 和 P19 (7.68 Å) 之间的紧密双链体内接触。后一种接触发生在 A 束的一端，但两个 Mg2+ 离子仅桥接磷酸盐跨过小沟，而没有穿透它。双工中的小凹槽是……
Much of what we know about B-form DNA stems from structural studies of the oligodeoxynucleotide CGCGAATTCGCG, the so-called Dickerson-Drew dodecamer (DDD). Its crystal structure provided the first detailed image of a right-handed DNA double helix. 1 Among the issues that were addressed based on this structure and those of related dodecamers are the interdependence of base sequence and structure, 2 backbone flexibility, 3 solvation, 4 bending and bendability, 3, 5 drug binding, 6 and the effects of packing forces7 and crystallization conditions8 on DNA structure. Intriguing features of the DDD duplex are the narrowness of the minor groove in the AATT region and the spine of water molecules in that groove. However, X-ray crystallography thus far failed to shed light on the effects of counterions, specifically mono-and divalent metal cations, on the structure of B-DNA. One of the reasons appears to be the limited resolution of DDD crystal structures (ca. 2.3 Å on average). On the basis of molecular dynamics (MD) simulations, it was suggested that Na+ ions can intrude electronegative “AT-pockets” in the minor groove and reside there with fractional occupancies. 9 NMR solution experiments of A-tract DNA provided evidence for the presence of Mn2+ ions in the minor groove. 10 The g1. 5 Å structures of the native DDD11 and a 12mer containing chemically modified thymidines12 prompted us to conduct a state-of-the-art crystallographic experiment with the goal to maximize the resolution of the DDD structure and learn more about the ionic environment of the duplex. Here, we report details of the DDD crystal structure at 1.1 Å resolution, the highest obtained so far for a B-DNA duplex.Among the factors that bring about this dramatically enhanced resolution are improvements over the last few years in the synthesis and purification of oligonucleotides. 13 However, modification of the original crystallization conditions, 14 proper freezing of crystals, and data collection at a third-generation synchrotron source15 are likely of more importance in this respect. Data collection and refinement17 statistics are summarized in Table 1. Three ordered Mg2+ ions are present per asymmetric unit, two hexahydrates (Mg1 and Mg3) and one pentahydrate complex (Mg2)(Figure 1; a, b, etc. designate symmetry mates). Mg1 is located in the major groove, close to one end of the duplex. 11, 12 The ion contacts the N7 and O6 edges of residues G2 and G22 from opposite strands via coordinated waters. It also bridges the O2P oxygens of P6c and P7c of an adjacent molecule and stabilizes the close interduplex contact between P2 and P7c (6.73 Å). This ion interaction likely causes the DDD duplex to asymmetrically kink into the major groove (Figure 2). 8 Additional close lateral interduplex contacts are seen between P20 and P12d (6.68 Å) and P10 and P18d (6.24 Å). Mg2 is directly coordinated to O1P of phosphate P19 and in addition forms a H bond to O1P of phosphate P12d via one of its water ligands. Similarly, Mg3 bridges oxygens O2P and O1P of phosphates P10 and P18d, respectively, through the same coordinated water (Figure 1). Mg2 stabilizes close contacts between phosphates P12 and P24 (5.59 Å) at both ends of the molecule. Thus, Mg2+ ions are located near the end-to-end overlaps between duplexes, a particular feature of the DDD lattice. 7 As shown in Figures 1 and 2, Mg2 and Mg3 also relieve a close intraduplex contact between phosphates P10 and P19 (7.68 Å). The latter contact occurs at one end of the A-tract, but the two Mg2+ ions only bridge the phosphates across the minor groove without penetrating it. The minor groove in the duplex is …