Polyamines favor DNA triplex formation at neutral pH.
Polyamines favor DNA triplex formation at neutral pH.
复制标题
多胺在中性 pH 值下有利于 DNA 三链体的形成。
作者：
Ken J. Hampel;Paul Crosson;Jeremy S. Lee
用四种不同的方法研究了三链DNA在多胺、精胺和亚精胺存在下的稳定性。首先，聚[d（TC）]、聚[d（GA）]的热变性分析表明，在低离子强度和pH 7下，3 μ M精胺足以引起所有双链体向三链体构象的歧化。高10倍浓度的亚精胺产生类似的效果。第二，在0.2M NaCl中在pH 5下测量歧化的动力学。添加500 μ M精胺使速率增加至少2倍。第三，在0.2 M NaCl中，在pH为5.8时发生三链体到三链体歧化的中点，但在500 μ M精胺存在下，这增加了近一个pH单位。第四，分子间三链体也可以在含有嘌呤嘧啶插入物的质粒中通过添加单链嘧啶形成。这在pH 7.2和25 mM离子强度下，在100 μ M精胺或亚精胺存在下很容易证明。然而，在0.2 M NaCl中，需要1 mM多胺。由于在真核细胞核中，多胺浓度在毫摩尔范围内，因此适当的嘌呤-嘧啶DNA序列可能有利于体内的三链体构象。
The stability of triplex DNA was investigated in the presence of the polyamines spermine and spermidine by four different techniques. First, thermal-denaturation analysis of poly[d(TC)].poly[d(GA)] showed that at low ionic strength and pH 7, 3 microM spermine was sufficient to cause dismutation of all of the duplex to the triplex conformation. A 10-fold higher concentration of spermidine produced a similar effect. Second, the kinetics of the dismutation were measured at pH 5 in 0.2 M NaCl. The addition of 500 microM spermine increased the rate by at least 2-fold. Third, in 0.2 M NaCl, the mid-point of the duplex-to-triplex dismutation occurred at a pH of 5.8, but this was increased by nearly one pH unit in the presence of 500 microM spermine. Fourth, intermolecular triplexes can also form in plasmids that contain purine.pyrimidine inserts by the addition of a single-stranded pyrimidine. This was readily demonstrated at pH 7.2 and 25 mM ionic strength in the presence of 100 microM spermine or spermidine. In 0.2 M NaCl, however, 1 mM polyamine is required. Since, in the eucaryotic nucleus, the polyamine concentration is in the millimolar range, then appropriate purine-pyrimidine DNA sequences may favor the triplex conformation in vivo.