Ferdous A, Watanabe H, Akaike T, Maruyama A
Department of Biomolecular Engineering, Faculty of Bioscience and Biotechnology, Tokyo Institute of Technology, 4259 Nagatsuta, Midori-ku, Yokohama 226-8501, Japan.
Nucleic Acids Res. 1998 Sep 1;26(17):3949-54. doi: 10.1093/nar/26.17.3949.
Triplex DNA formation involving unmodified triplex-forming oligonucleotides (TFOs) is very unstable under physiological conditions. Here, we report a novel strategy to stabilize both purine and pyrimidine motif triplex DNA within the rat alpha1 (I) collagen gene promoter under physiologically relevant conditions by a poly(L-lysine)- graft -dextran copolymer. Using an in vitro electrophoretic mobility shift assay, we show that the copolymer almost completely abrogates the inhibitory effects of physiological concentrations of monovalent cations, particularly potassium ion (K+), on purine motif triplex formation involving very low concentrations of an unmodified guanine-rich TFO. Of importance, pH dependency in pyrimidine motif triplex formation involving an unmodified cytosine-rich TFO is also significantly overcome by the copolymer. Finally, the triplex-stabilizing efficiency of the copolymer is remarkably higher than that of other oligocations, like spermine and spermidine. We suggest that the ability of the graft copolymer to stabilize triplex DNA under physiologically relevant pH and salt concentrations will be a cue for further progress in the antigene strategy.
涉及未修饰三链体形成寡核苷酸（TFOs）的三链体DNA形成在生理条件下非常不稳定。在此，我们报告了一种新策略，即在生理相关条件下，通过聚（L-赖氨酸）-接枝-葡聚糖共聚物来稳定大鼠α1（I）胶原基因启动子内的嘌呤和嘧啶基序三链体DNA。使用体外电泳迁移率变动分析，我们表明该共聚物几乎完全消除了生理浓度的单价阳离子，特别是钾离子（K+），对涉及极低浓度未修饰富含鸟嘌呤TFO的嘌呤基序三链体形成的抑制作用。重要的是，该共聚物也显著克服了涉及未修饰富含胞嘧啶TFO的嘧啶基序三链体形成中的pH依赖性。最后，该共聚物的三链体稳定效率明显高于其他寡阳离子，如精胺和亚精胺。我们认为，接枝共聚物在生理相关的pH和盐浓度下稳定三链体DNA的能力将是反基因策略进一步发展的一个线索。
