Rüba Eva, Hart Jonathan R, Barton Jacqueline K
Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.
Inorg Chem. 2004 Jul 26;43(15):4570-8. doi: 10.1021/ic0499291.
Here we report the synthesis of luminescent ruthenium complexes that bind DNA base pair mismatches. [Ru(bpy)2(tpqp)]Cl2 (tpqp = 7,8,13,14-tetrahydro-6-phenylquino[8,7-k][1,8]phenanthroline), [Ru(bpy)2(pqp)]Cl2 (pqp = 6-phenylquino[8,7-k][1,8]phenanthroline), and [Ru(bpy)2(tactp)]Cl2 [tactp = 4,5,9,18-tetraazachryseno[9,10-b]triphenylene] have been synthesized, and their spectroscopic properties in the absence and presence of DNA have been examined. While [Ru(bpy)2(pqp)]2+ shows no detectable luminescence, [Ru(bpy)2(tpqp)]2+ is luminescent in the absence and presence of DNA with an excited-state lifetime of 10 ns and a quantum yield of 0.002. Although no increase in emission intensity is associated with binding to mismatch-containing DNA, luminescence quenching experiments and measurements of steady-state fluorescence polarization provide evidence for preferential binding to oligonucleotides containing a CC mismatch. Furthermore, by marking the site of binding through singlet oxygen sensitized damage, the complex has been shown to target a CC mismatch site directly with a specific binding affinity, Kb = 4 x 10(6) M(-1). [Ru(bpy)2(tactp)]2+, an analogue of [Ru(bpy)2(dppz)]2+ containing a bulky intercalating ligand, is luminescent in aqueous solution at micromolar concentrations and exhibits a 12-fold enhancement in luminescence in the presence of DNA. The complex, however, tends to aggregate in aqueous solution; we find a dimerization constant of 9.8 x 10(5) M(-1). Again, by singlet oxygen sensitization it is apparent that [Ru(bpy)2(tactp)]2+ binds preferentially to a CC mismatch; using a DNase I footprinting assay, a binding constant to a CC mismatch of 8 x 10(5) M(-1) is found. Hence results with these novel luminescent complexes support the concept of using a structurally demanding ligand to obtain selectivity in targeting single base mismatches in DNA. The challenge is coupling the differential binding we can obtain to differential luminescence.
在此,我们报告了可结合DNA碱基对错配的发光钌配合物的合成。已合成了[Ru(bpy)₂(tpqp)]Cl₂(tpqp = 7,8,13,14 - 四氢 - 6 - 苯基喹啉并[8,7 - k][1,8]菲咯啉)、[Ru(bpy)₂(pqp)]Cl₂(pqp = 6 - 苯基喹啉并[8,7 - k][1,8]菲咯啉)和[Ru(bpy)₂(tactp)]Cl₂[tactp = 4,5,9,18 - 四氮杂蒽并[9,10 - b]三苯撑],并研究了它们在不存在和存在DNA时的光谱性质。虽然[Ru(bpy)₂(pqp)]²⁺未显示出可检测到的发光,但[Ru(bpy)₂(tpqp)]²⁺在不存在和存在DNA时均发光,其激发态寿命为10 ns,量子产率为0.002。尽管与含错配的DNA结合未伴随发射强度增加,但发光猝灭实验和稳态荧光偏振测量为优先结合含CC错配的寡核苷酸提供了证据。此外,通过单线态氧敏化损伤标记结合位点,已证明该配合物以特定结合亲和力Kb = 4×10⁶ M⁻¹直接靶向CC错配位点。[Ru(bpy)₂(tactp)]²⁺是含庞大嵌入配体的[Ru(bpy)₂(dppz)]²⁺的类似物,在微摩尔浓度的水溶液中发光,且在存在DNA时发光增强12倍。然而,该配合物在水溶液中倾向于聚集;我们测得其二聚化常数为9.8×10⁵ M⁻¹。同样,通过单线态氧敏化可知[Ru(bpy)₂(tactp)]²⁺优先结合CC错配;使用DNase I足迹分析,发现其与CC错配的结合常数为8×10⁵ M⁻¹。因此,这些新型发光配合物的结果支持了使用结构上有要求的配体来获得靶向DNA中单碱基错配选择性的概念。挑战在于将我们能获得的差异结合与差异发光相耦合。