Trapane T L, Hogrefe R I, Reynolds M A, Kan L S, Ts'o P O
Department of Biochemistry, School of Hygiene and Public Health, Johns Hopkins University, Baltimore, Maryland 21205, USA.
Biochemistry. 1996 Apr 30;35(17):5495-508. doi: 10.1021/bi960070b.
We have investigated the role of purines in interstrand complex formation with regard to substitution of the negatively-charged, phosphodiester backbone by a nonionic, internucleoside linkage. Using the purine oligomer, d(AG)8, its methylphosphonate analog, d(AG)8, and the complementary pyrimidine oligomer, d(CT)8, as a model system, the stoichiometry, conformation, and stability of complexes formed at pH 8 were studied by spectroscopic and electrophoretic methods. When there is only one oligomer species in solution, d(AG)8 behaves as a single-stranded molecule. In contrast, the d(AG)8 oligomer readily forms an intermolecular self-complex, particularly in the presence of magnesium ion. Using either purine oligomer, duplexes can form with the d(CT)8 strand which differ in terms of their conformation and in the dependence of their thermal stability on sodium and magnesium ions. All studies show that a stable triplex forms with a 1:2 d(CT)8:d(AG)8 stoichiometry which does not require high concentrations of sodium or magnesium ions. Triplex formation between the d(CT)8 strand and two d(AG)8 strands was not observed. Native gel electrophoresis suggests that a 1:1:1 d(CT)8:d(AG)8:d(AG)8 complex may be formed. In regard to triplex formation, the advantage of the methylphosphonate backbone on the purine strand is clearly demonstrated.
我们研究了嘌呤在链间复合物形成中的作用，涉及用非离子型核苷间连接取代带负电荷的磷酸二酯主链。以嘌呤寡聚物d(AG)8、其甲基膦酸酯类似物d(AG)8以及互补嘧啶寡聚物d(CT)8作为模型系统，通过光谱和电泳方法研究了在pH 8时形成的复合物的化学计量、构象和稳定性。当溶液中只有一种寡聚物种时，d(AG)8表现为单链分子。相比之下，d(AG)8寡聚物很容易形成分子间自复合物，特别是在镁离子存在的情况下。使用任何一种嘌呤寡聚物，都可以与d(CT)8链形成双链体，它们在构象以及热稳定性对钠和镁离子的依赖性方面存在差异。所有研究表明，以1:2的d(CT)8:d(AG)8化学计量形成稳定的三链体，这不需要高浓度的钠或镁离子。未观察到d(CT)8链与两条d(AG)8链之间形成三链体。天然凝胶电泳表明可能形成1:1:1的d(CT)8:d(AG)8:d(AG)8复合物。关于三链体的形成，可以清楚地证明嘌呤链上甲基膦酸酯主链的优势。
