Sato Yusuke, Seino Takehiro, Nishizawa Seiichi, Teramae Norio
Department of Chemistry, Graduate School of Science, Tohoku University, Aoba-ku, Sendai 980-8578, Japan.
Nucleic Acids Symp Ser (Oxf). 2006(50):219-20. doi: 10.1093/nass/nrl109.
We here report on the thermodynamics of the hydrogen bond-mediated binding of 2-amino-7-methyl-1,8-naphthyridine (AMND) to a cytosine base opposite an abasic site (AP site) in a 21-meric DNA duplex (5'-GCA GCT CCC GXG GTC TCC TCG-3'/3'-CGT CGA GGG CCC CAG AGG AGC-5', X= AP site, C = target). The examination by fluorescence titration experiments shows a 1:1 binding constant of 2.7x10(6) M(-1) at 20 degrees C in solutions containing 110 mM Na(+) (pH 7.0). From the analysis of salt dependence of binding constants, polyelectrolyte (DeltaG(pe)) and non-polyelectrolyte (DeltaG(t)) contributions are calculated as -1.7 kcal/mol and -6.9 kcal/mol, respectively, at 110 mM Na(+) concentration. The binding enthalpy determined by isothermal titration calorimetry (ITC) is -18.5 kcal/mol in 110 mM Na(+) at 20 degrees C. We discuss these results with a view towards further development of our ligand-based fluorescence assay for SNPs (single nucleotide polymorphisms) typing.
我们在此报告2-氨基-7-甲基-1,8-萘啶（AMND）与21聚体DNA双链体（5'-GCA GCT CCC GXG GTC TCC TCG-3'/3'-CGT CGA GGG CCC CAG AGG AGC-5'，X = 无碱基位点，C = 靶标）中与无碱基位点（AP位点）相对的胞嘧啶碱基通过氢键介导的结合的热力学。荧光滴定实验表明，在含110 mM Na⁺（pH 7.0）的溶液中，20℃时结合常数为1:1，为2.7×10⁶ M⁻¹。通过对结合常数的盐依赖性分析，在110 mM Na⁺浓度下，聚电解质（ΔG(pe)）和非聚电解质（ΔG(t)）贡献分别计算为-1.7 kcal/mol和-6.9 kcal/mol。在20℃下，110 mM Na⁺中通过等温滴定量热法（ITC）测定的结合焓为-18.5 kcal/mol。我们讨论这些结果，以期进一步开发基于配体的单核苷酸多态性（SNP）分型荧光测定法。
