Pilch D S, Brousseau R, Shafer R H
Graduate Group in Biophysics, School of Pharmacy, University of California, San Francisco 94143.
Nucleic Acids Res. 1990 Oct 11;18(19):5743-50. doi: 10.1093/nar/18.19.5743.
We have stabilized the d(A)10.2d(T)10 and d(C+LT4C+3).d(G3A4G3).d(C3T4C3) triple helices with either NaCl or MgCl2 at pH 5.5. UV mixing curves demonstrate a 1:2 stoichiometry of purine to pyrimidine strands under the appropriate conditions of pH and ionic strength. Circular dichroic titrations suggest a possible sequence-independent spectral signature for triplex formation. Thermal denaturation profiles indicate the initial loss of the third strand followed by dissociation of the underlying duplex with increasing temperature. Depending on the base sequence and ionic conditions, the binding affinity of the third strand for the duplex at 25 degrees C is two to five orders of magnitude lower than that of the two strands forming the duplex. Thermodynamic parameters for triplex formation were determined for both sequences in the presence of 50 mM MgCl2 and/or 2.0 M NaCl. Hoogsteen base pairs are 0.22-0.64 kcal/mole less stable than Watson-Crick base pairs, depending on ionic conditions and base composition. C+.G and T.A Hoogsteen base pairs appear to have similar stability in the presence of Mg2+ ions at low pH.
我们已在pH 5.5条件下，使用NaCl或MgCl2使d(A)10.2d(T)10以及d(C+LT4C+3).d(G3A4G3).d(C3T4C3)三链螺旋结构得以稳定。紫外混合曲线表明，在合适的pH和离子强度条件下，嘌呤链与嘧啶链的化学计量比为1:2。圆二色滴定表明三链形成可能存在与序列无关的光谱特征。热变性曲线表明，随着温度升高，首先是第三条链的初始丢失，随后是下层双链的解离。根据碱基序列和离子条件，第三条链在25℃时对双链的结合亲和力比形成双链的两条链之间的结合亲和力低两到五个数量级。在50 mM MgCl2和/或2.0 M NaCl存在的情况下，测定了两种序列形成三链的热力学参数。根据离子条件和碱基组成，Hoogsteen碱基对比沃森-克里克碱基对的稳定性低0.22 - 0.64千卡/摩尔。在低pH条件下存在Mg2+离子时，C+.G和T.A Hoogsteen碱基对似乎具有相似的稳定性。
