【摘要】 为考察小分子配基与不同核酸结构的结合机理,发展新的核酸探针分子,合成了一种新型一次甲基不对称菁染料(MTP).吸收光谱、荧光光谱及圆二色光谱研究结果表明,MTP可与平行和混合平行G-四链体DNA(如c-myc和22AGK+)较强地结合,并引起130~180倍的荧光增强;其与单/双链DNA作用较弱,导致40~60倍荧光增强;而与反平行G-四链体DNA(如TBA和22AGNa+)的作用最弱,只引起15~25倍荧光增强;以上结果表明MTP可作为荧光探针分子用于区别不同结构的核酸分子.结合机理研究表明,平行和混合平行G-四链体DNA优先通过沟槽结合模式结合1分子MTP,再通过末端堆积模式结合另1分子MTP.更多还原

【Abstract】 Cyanine dyes have been widely used as fluorescent probes for nucleic acids. A well-known monomethine cyanine dye,thiazole orange(TO),has been reported to bind various forms of nucleic acids,such as RNA,duplex DNA,triplex DNA and especially G-quadruplex DNA. As a nucleic acid light-up probe,TO has been used in a variety of applications. The binding modes between TO and G-quadruplex DNA are complex,may include end-stacking,groove binding and external stacking,the predominant binding mode is reported to be end-stacking. In order to further understand the interaction of monomethine cyanines with different nucleic acid forms and develop new probes,we synthesized a new analogue of TO,1,2-dimethyl-6-{[3-methylbenzo[d]thiazol-2(3H)-ylidene]methyl} pyridine-1-ium( MTP) and investigated its interaction with different DNA forms. The interaction of MTP with c-myc( parallel G-quadruplex),22AGK+(mixed-type G-quadruplex),ss /dsDNA(single /double-stranded DNA) caused notable red-shift and hypochromicity of the absorption spectrum of MTP. MTP exhibited almost no fluorescence in aqueous buffer condition. However,the interaction of MTP with DNA resulted in great enhancement of MTP fluorescence,approximately 130—180-fold for c-myc or 22AGK+,40—60-fold for single/double-stranded(ss/ds) DNA and 15—25-fold for TBA and 22AGNa+(antiparallel G-quadruplexes). The apparent dissociation constants(Kd) of MTP and different DNA were in the range of 4. 0—17 !mol /L. The fluorescence enhancement was corrected with the binding affinity of MTP and different DNA forms. These results suggest that MTP can be used as a fluorescent probe to distinguish different forms of nucleic acids. The binding stoichiometry showed that two molecules of MTP bound to one molecule of c-myc or 22AGK+. The induced CD spectroscopy and G-quadruplex /hemin peroxidase inhibition experiment suggested that the first MTP bound to c-myc through the groove binding model and the second MTP bound to c-myc through the end-stacking model. Compared with TO,MTP has a smaller size;it showed different absorption spectral changes upon binding to DNA and exhibited different fluorescence responses to 22AGK+and 22AGNa+,which may suggest the different binding modes of TO and MTP to G-quadruplexes. These results provide important information for the design of the ligands and probes for nucleic acids.更多还原