2-aminopurine fluorescence studies of base stacking interactions at abasic sites in DNA: metal-ion and base sequence effects
2-aminopurine fluorescence studies of base stacking interactions at abasic sites in DNA: metal-ion and base sequence effects
复制标题
DOI：
10.1093/nar/26.16.3837
复制
发表时间：
1998-08-15
影响因子：
14.9
通讯作者：
Stivers, JT
中科院分区：
文献类型：
--
作者：
Stivers, JT
利用荧光核苷酸探针2-氨基嘌呤（2-AP）检测了10种不同DNA双链中碱基（AB）位点周围碱基堆叠相互作用的金属离子和序列依赖性变化，这些碱基与碱基（AB）位点相对（AB- ap (opp)）或与碱基（AB- ap (adj)）相邻。对这些AB双工及其母体双工的荧光发射和激发光谱的详细研究表明，AB- ap （opp）在相应的正常双工中明显少于2-AP的堆叠。一般来说，AB链上的AB- ap （adj）是堆叠的，而相反链上的AB- ap （adj）表现出不稳定的堆叠相互作用。结果还表明，二价阳离子与AB- ap （opp）的结合有助于AB- ap （adj）的基层相互作用的不稳定，但对AB- ap （adj）的堆叠相互作用影响很小或没有影响。与这些结果一致，AB- ap （opp）的荧光对外部添加猝灭剂的敏感性是母体正常双相的18-30倍。当尿嘧啶DNA糖基酶在2.5 mM MgCl2存在下与AB-AP（opp）结合时，观察到荧光下降3倍（K-d = 400 +/- 90 nM），表明未堆叠的2-AP（opp）在结合后变得更加堆叠。在这些荧光研究的基础上，提出了这些AB位点的局部碱基堆叠相互作用模型。
Metal-ion and sequence dependent changes in the stacking interactions of bases surrounding abasic (AB) sites in 10 different DNA duplexes were examined by incorporating the fluorescent nucleotide probe 2-aminopurine (2-AP), opposite to the site (AB-AP(opp)) or adjacent to the site (AB-AP(adj)) on either strand. A detailed study of the fluorescence emission and excitation spectra of these AB duplexes and their corresponding parent duplexes indicates that AB-AP(opp) is significantly less stacked than 2-AP in the corresponding normal duplex. In general, AB-AP(adj) on the AB strand is stacked, but AB-AP(adj) on the opposite strand shows destabilized stacking interactions. The results also indicate that divalent cation binding to the AB duplexes contributes to destabilizaton of the base stacking interactions of AB-AP(opp), but has little or no effect on the stacking interactions of AB-AP(adj), Consistent with these results, the fluorescence of AB-AP(opp) is 18-30-fold more sensitive to an externally added quenching agent than the parent normal duplex. When uracil DNA glycosylase binds to AB-AP(opp) in the presence of 2.5 mM MgCl2, a 3-fold decrease in fluorescence is observed (K-d = 400 +/- 90 nM) indicating that the unstacked 2-AP(opp) becomes more stacked upon binding. On the basis of these fluorescence studies a model for the local base stacking interactions at these AB sites is proposed.