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对来自突变热点序列的含凸起DNA寡核苷酸的质子核磁共振研究。

Proton nuclear magnetic resonance studies on bulge-containing DNA oligonucleotides from a mutational hot-spot sequence.

作者信息

Woodson S A, Crothers D M

出版信息

Biochemistry. 1987 Feb 10;26(3):904-12. doi: 10.1021/bi00377a035.

DOI:10.1021/bi00377a035
PMID:3567151
Abstract

A series of bulge-containing and normal double-helical synthetic oligodeoxyribonucleotides, of sequence corresponding to a frame-shift mutational hot spot in the lambda C1 gene, are compared by proton magnetic resonance spectroscopy at 500 MHz. The imino proton resonances of d(GATGGGCAG).d(CTGCCCCATC), d(GATGGGCAG).d(CTGCCCCATC), and d(GATGGGCAG).d(CTGACCCATC) are assigned by one-dimensional nuclear Overhauser effect spectroscopy. Nonselective T1 inversion-recovery experiments are used to determine exchangeable proton lifetimes and to compare helix stability and dynamics of the three duplexes. An extra adenosine flanking the internal G.C base pairs has a strongly localized effect on helix stability, but the destabilizing effect of an extra cytidine in a C tract is delocalized over the entire G.C run. These data lead to the conclusion that the position of the bulge migrates along the run in the fast-exchange limit on the NMR time scale. Rapid migration of the bulge defect in homopolymeric sequences may help rationalize both frame-shift mutagenesis and translational frame shifting. We estimate that the unfavorable free energy of a localized bulge defect is 2.9-3.2 kcal/mol, in good agreement with earlier estimates for RNA helices.

摘要

通过500兆赫的质子磁共振光谱法，比较了一系列含有凸起和正常双螺旋的合成寡脱氧核糖核苷酸，其序列对应于λ C1基因中的一个移码突变热点。d(GATGGGCAG).d(CTGCCCCATC)、d(GATGGGCAG).d(CTGCCCCATC)和d(GATGGGCAG).d(CTGACCCATC)的亚氨基质子共振通过一维核Overhauser效应光谱法进行归属。非选择性T1反转恢复实验用于确定可交换质子寿命，并比较三种双链体的螺旋稳定性和动力学。内部G.C碱基对侧翼的一个额外腺苷对螺旋稳定性有强烈的局部影响，但C序列中一个额外胞苷的去稳定作用在整个G.C序列上是分散的。这些数据得出结论，在核磁共振时间尺度的快速交换极限下，凸起的位置沿序列迁移。同聚物序列中凸起缺陷的快速迁移可能有助于解释移码诱变和平移移码现象。我们估计局部凸起缺陷的不利自由能为2.9 - 3.2千卡/摩尔，与早期对RNA螺旋的估计非常吻合。

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