Conformer Analysis of a Biological G-Quadruplex Element in the Human c-MYC Promoter by Native Polyacrylamide Gel Electrophoresis
Conformer Analysis of a Biological G-Quadruplex Element in the Human c-MYC Promoter by Native Polyacrylamide Gel Electrophoresis
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通过天然聚丙烯酰胺凝胶电泳对人 c-MYC 启动子中的生物 G-四链体元件进行构象分析
DOI：
10.1007/s41664-021-00181-0
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发表时间：
2021
影响因子：
4.7
通讯作者：
H. Zuo
中科院分区：
文献类型：
--
作者：
Xiao;Hong;Liang Fang;H. Zuo
g -四聚体经历复杂的折叠和构象交换。g -四相化合物的稳定性受序列、缓冲液和温度的影响较大。突变分析与核磁共振（NMR）、x射线晶体学和圆二色光谱已被证明是g -四重体结构分析的有力方法。在此，我们利用DNA序列突变和天然聚丙烯酰胺凝胶电泳研究了人类c-MYC启动子中发现的g -四重体模型分子Pu18的拓扑结构和构象。c-MYC Pu18序列中不参与g四分体形成的鸟嘌呤（G6、G9和G18）突变为胸腺嘧啶或腺嘌呤。我们筛选了Pu18凝胶电泳的缓冲液和温度。凝胶电泳显示，在100 mM K+缓冲液中，c-MYC Pu18的4个构象中有2个被分解，这与前人研究中1H NMR谱确定的构象一致。该技术操作简单，成本低，有望在核磁共振、x射线晶体学和圆二色光谱等其他分析方法的帮助下，成为一种通用的方法，有助于发现更多尚未确定的g -四重折叠和功能。
The G-quadruplexes undergo complex folding and conformation exchanges. G-quadruplex stability is substantially influenced by sequence, buffer and temperature. Mutational analysis together with nuclear magnetic resonance spectroscopy (NMR), X-ray crystallography and circular dichroism spectroscopy has been proved to be a powerful approach for G-quadruplex structural analysis. Herein, we used DNA sequence mutation and native polyacrylamide gel electrophoresis to investigate the topology and conformations of a G-quadruplex model molecule Pu18 found in the human c-MYC promoter. The guanines (G6, G9 or G18) which were not contributable to G-tetrad formation in c-MYC Pu18 sequence were mutated to thymine or adenine. We screened the buffer and temperature of gel electrophoresis for Pu18. Gel electrophoresis showed that two of the four conformers of c-MYC Pu18 in 100 mM K+ buffer were resolved, which was in accordance with the conformations as determined by the 1H NMR spectra in previous studies. This technique is expected as a general methodology for its easy operation and low cost to facilitate uncovering more yet unidentified G-quadruplex folds and functions, with the assistance of other analytical methods like NMR, X-ray crystallography and circular dichroism spectroscopy.
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影响因子：
14.9
作者：
Rhodes D;Lipps HJ
通讯作者：
Lipps HJ
影响因子：
15
作者：
Seenisamy, J;Rezler, EM;Hurley, LH
通讯作者：
Hurley, LH
影响因子：
2.9
作者：
Ambrus, A;Chen, D;Yang, DZ
通讯作者：
Yang, DZ
影响因子：
--
作者：
He Y;Tian Y;Mao C
通讯作者：
Mao C