Ray A, Kumar G S, Das S, Maiti M
Biophysical Chemistry Laboratory, Indian Institute of Chemical Biology, 4 Raja S.C. Mullick Road, Calcutta 700032, India.
Biochemistry. 1999 May 11;38(19):6239-47. doi: 10.1021/bi982128n.
The interaction of aristololactam-beta-D-glucoside (ADG), a DNA intercalating alkaloid, with the DNA triplexes, poly(dT). poly(dA)xpoly(dT) and poly(dC).poly(dG)xpoly(dC+), and the RNA triplex poly(rU).poly(rA)xpoly(rU) was investigated by circular dichroic, UV melting profile, spectrophotometric, and spectrofluorimetric techniques. Comparative interaction with the corresponding Watson-Crick duplexes has also been examined under identical experimental conditions. Triplex formation has been confirmed from biphasic thermal melting profiles and analysis of temperature-dependent circular dichroic measurements. The binding of ADG to triplexes and duplexes is characterized by the typical hypochromic and bathochromic effects in the absorption spectrum, quenching of steady-state fluorescence intensity, a decrease in fluorescence quantum yield, an increase or decrease of thermal melting temperatures, and perturbation in the circular dichroic spectrum. Scatchard analysis indicates that ADG binds both to the triplexes and the duplexes in a noncooperative manner. Binding parameters obtained from spectrophotometric measurements are best fit by the neighbor exclusion model. The binding affinity of ADG to the DNA triplexes is substantially stronger than to the RNA triplex. Thermal melting study further indicates that ADG stabilizes the Hoogsteen base-paired third strand of the DNA triplexes whereas it destabilizes the same strand of RNA triplex but stabilizes its Watson-Crick strands. Comparative data reveal that ADG exhibits a stronger binding to the triple helical structures than to the respective double helical structures.
采用圆二色性、紫外熔解曲线、分光光度法和荧光光谱法等技术，研究了DNA嵌入生物碱马兜铃内酰胺-β-D-葡萄糖苷（ADG）与DNA三链体poly(dT).poly(dA)xpoly(dT)、poly(dC).poly(dG)xpoly(dC+)以及RNA三链体poly(rU).poly(rA)xpoly(rU)之间的相互作用。在相同实验条件下，还研究了ADG与相应的沃森-克里克双链体的相互作用。通过双相热熔解曲线和温度依赖性圆二色性测量分析，证实了三链体的形成。ADG与三链体和双链体的结合表现为吸收光谱中典型的减色和红移效应、稳态荧光强度猝灭、荧光量子产率降低、热熔解温度升高或降低以及圆二色光谱扰动。Scatchard分析表明，ADG以非协同方式与三链体和双链体结合。分光光度测量获得的结合参数最适合邻位排斥模型。ADG与DNA三链体的结合亲和力明显强于与RNA三链体的结合亲和力。热熔解研究进一步表明，ADG使DNA三链体的Hoogsteen碱基配对第三条链稳定，而使RNA三链体的同一条链不稳定，但使其沃森-克里克链稳定。比较数据表明，ADG对三螺旋结构的结合比对相应双螺旋结构的结合更强。
