单分子荧光成像研究凝血酶核酸适体的折叠

Single Molecule Fluorescence Imaging of Thrombin Aptamer Folding

通过对固定在表面的TMR标记凝血酶核酸适体进行单分子荧光成像,在单分子水平上研究了凝血酶核酸适体的折叠.在有K+存在的条件下,核酸适体分子与K+结合后发生折叠,形成G四分体结构,使得TMR靠近富含鸟嘌呤的G四分体,并与鸟嘌呤发生电子转移,从而导致TMR荧光强度降低.根据TMR的单分子荧光强度观察到不同K+浓度下核酸适体在折叠和无规卷曲两种状态下的分布.结果表明,可利用电子转移引起的荧光强度变化在单分子水平上研究核酸适体构象变化,这一新方法的建立是对常用的单分子荧光共振能量转移(FRET)法的重要补充.

The folding of the thrombin DNA aptamer was studied with single molecule fluorescence imaging. The aptamer was singly labeled with TMR and immobilized on the glass surface. In the presence of K^＋ , the folding of the aptamer resulted in the formation of G-quartets, thus brought TMR close to the guanosines. This led to the fluorescence decrease of TMR through the electron transfer between the dye and guanosines. From the fluorescence intensity of individual TMR, the distribution of aptamer molecules between the folded and random coil conformations was monitored at different concentrations of K^＋. The results show a new approach for the investigation of aptamer conformational change by electron transfer induced fluorescence quenching, which is complementary to the commonly used fluorescence resonance energy transfer in single molecule study.