Sequence and environmental effects on the self-assembly of DNA oligomers possessing G(x)T(2)G(y) segments
Sequence and environmental effects on the self-assembly of DNA oligomers possessing G(x)T(2)G(y) segments
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DOI：
10.1021/bi960979u
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发表时间：
1996-08-13
期刊：
影响因子：
2.9
通讯作者：
Sheardy, RD
中科院分区：
文献类型：
--
作者：
Marotta, SP;Tamburri, PA;Sheardy, RD
众所周知，具有连续鸟嘌呤碱基的DNA寡聚体可以通过形成四链物种而呈现非Watson-Crick类型的结构。这些四链结构的结构高度依赖于DNA的序列和制备DNA的条件(例如，缓冲液、pH、离子强度、阳离子存在和温度)。本实验室先前已经报道了序列C(4)T(4)G(4)T(1-4)G(4)的DNA低聚物自组装成多链高分子量物种[DAL，T-Y，Marotta，S.P.，&Seardy，R.D.(1995)生化34,3655-3662]，以进一步研究序列和环境对具有G(X)T(2)G(Y)片段的DNA低聚物自组装的影响，其中x=1，3或4，并且y=2-5，合成了许多这样的低聚物。在标准磷酸盐缓冲液(10 mM磷酸盐，pH 7.0)和添加不同浓度的氯化钠、氯化钾和/或氯化镁中制备DNA样品，以评价阳离子及其浓度对DNA低聚体自组装的影响。用非变性聚丙烯酰胺凝胶电泳法和圆二色谱研究了这些低聚物的自组装情况。低聚物在100 mM K+或50 mM Na+和50 mM K+中的电泳表明，这些低聚物形成了一个或两个分子物种。相反，在100 mM K+和20 mM Mg2+存在的情况下对这些低聚物进行电泳，可以得到多条高相对分子质量的梯形条带，表明多标准DNA结构的形成。结果表明，镁离子的存在以及G(Y)末端四个或更多碱基的存在有利于自组装成高相对分子质量物种。这些结果还表明，具有相似序列的端粒DNA的结构可能非常不寻常。
It is well-known that DNA oligomers possessing contiguous guanine bases call assume non-Watson-Crick type structures through the formation of four-stranded species. The architecture of these four-stranded structures is highly dependent upon the sequence of the DNA and the conditions (e.g., buffer, pH, ionic strength, cations present, and temperature) under which the DNA is prepared. This lab has previously reported the self-assembly of DNA oligomers of sequence C(4)T(4)G(4)T(1-4)G(4) into multistranded high molecular weight: species [Dal, T.-Y., Marotta, S. P., & Sheardy, R. D. (1995) Biochemistry 34, 3655-3662], In order to further investigate the sequence and environmental effects on the self-assembly of DNA oligomers possessing G(x)T(2)G(y) (where x = 1, 3, or 4 and y = 2-5) segments, the synthesis of a number of such oligomers was undertaken. DNA samples were prepared in standard phosphate buffer (10 mM phosphate, pH 7.0) and NaCl, KCI and/or MgCl2 added to different concentrations in order to evaluate the influence of the cations and their concentrations on the self-assembly of the DNA oligomers. The self-assembly of these oligomers was monitored by nondenaturing polyacrylamide gel electrophoresis and circular dichroism studies. Electrophoresis of the oligomers in either 100 mM K+ or 50 mM Na+ with 50 mM K+ indicated the formation of one or two molecular species for these oligomers. In contrast, electrophoresis of these oligomers in the presence of both 100 mM K+ and 20 mM Mg2+ give a ladder of multiple bands of high molecular weight indicative of multistanded DNA structure formation. The results presented here indicate that self-assembly into high molecular weight species is favored by the presence of Mg2+ as well as the presence of four or more bases in the terminal G(y) segment. These results also suggest that the structure of telomeric DNA, which possesses similar sequences, may be quite unusual.