Kinetics and mechanism of K+- and Na+-induced folding of models of human telomeric DNA into G-quadruplex structures.
Kinetics and mechanism of K+- and Na+-induced folding of models of human telomeric DNA into G-quadruplex structures.
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DOI：
10.1093/nar/gkn379
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发表时间：
2008-07
影响因子：
14.9
通讯作者：
Chaires, Jonathan B.
中科院分区：
文献类型：
--
作者：
Gray, Robert D.;Chaires, Jonathan B.
用KCl和NaCl平衡滴定法和多波长停流动力学方法研究了3种模型人端粒寡核苷酸d[AGGG（TTAGGG）3]、d[TTGGG（TTAGGG）3A]和d[TTGGG（TTAGGG）3]在阳离子诱导下折叠成四链体的过程。阳离子结合是协同的，希尔系数在K+中为1.5-2.2，在Na+中为2.4-2.9，半饱和浓度K+为0.5-1 mM，Na+为4-13 mM，这取决于寡核苷酸序列。在25°C下在50 mM KCl中的寡核苷酸折叠由单指数过程组成，弛豫时间τ为20-60 ms，取决于序列。相比之下，在100 mM NaCl中的折叠由三个指数组成，τ值为40-85 ms，250-950 ms和1.5-10.5 s。随着阳离子浓度的增加，折叠速率常数接近极限值;此外，在15-45°C范围内，折叠速率随着温度的升高而降低。两者合计，这些结果表明，富含G的寡核苷酸折叠成四链体结构通过动力学显著的中间体进行。这些中间体可能包括反平行发夹结构，在快速平衡，有序结构较少。发夹随后可以形成新生的G-四联体，其通过H-键合和阳离子结合而稳定，随后是相对缓慢的链重排以形成最终的完全折叠的拓扑结构。较少的动力学中间体与K+比Na+明显，这表明在K+溶液中的折叠途径更简单。
Cation-induced folding into quadruplex structures for three model human telomeric oligonucleotides, d[AGGG(TTAGGG)3], d[TTGGG(TTAGGG)3A] and d[TTGGG(TTAGGG)3], was characterized by equilibrium titrations with KCl and NaCl and by multiwavelength stopped flow kinetics. Cation binding was cooperative with Hill coefficients of 1.5–2.2 in K+ and 2.4–2.9 in Na+ with half-saturation concentrations of 0.5–1 mM for K+ and 4–13 mM for Na+ depending on the oligonucleotide sequence. Oligonucleotide folding in 50 mM KCl at 25°C consisted of single exponential processes with relaxation times τ of 20–60 ms depending on the sequence. In contrast, folding in100 mM NaCl consisted of three exponentials with τ-values of 40–85 ms, 250–950 ms and 1.5–10.5 s. The folding rate constants approached limiting values with increasing cation concentration; in addition, the rates of folding decreased with increasing temperature over the range 15–45°C. Taken together, these results suggest that folding of G-rich oligonucleotides into quadruplex structures proceeds via kinetically significant intermediates. These intermediates may consist of antiparallel hairpins in rapid equilibrium with less ordered structures. The hairpins may subsequently form nascent G-quartets stabilized by H-bonding and cation binding followed by relatively slow strand rearrangements to form the final completely folded topologies. Fewer kinetic intermediates were evident with K+ than Na+, suggesting a simpler folding pathway in K+ solutions.
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