Verdian-Doghaei A, Housaindokht M R, Bozorgmehr M R, Abnous K
a Biophysical Chemistry Laboratory, Department of Chemistry, Faculty of Science , Ferdowsi University of Mashhad , Mashhad , Iran.
J Biomol Struct Dyn. 2015;33(6):1153-63. doi: 10.1080/07391102.2014.935482. Epub 2014 Jul 11.
Guanine-rich sequences can form the G-quadruplex structure in the presence of specific metal ions. Here, circular dichroism, UV-vis absorption, fluorescence, and molecular dynamics simulation studies revealed that insulin-binding aptamer (IBA) could form an intramolecular G-quadruplex structure after binding K(+). Circular dichroism (CD) spectra demonstrated that IBA could fold into a parallel G-quadruplex with a strong positive peak at 263 nm. Analysis of equilibrium titration data revealed that cation binding was cooperative with the Hill coefficient of 2.01 in K(+) and 1.90 in Na(+). Thermal denaturation assays indicated that K(+)-induced G-quadruplex is more stable than Na(+)-induced structure. Folding of IBA into G-quadruplex leading to the contact quenching occurs as a result of the formation of a nonfluorescent complex between donor and acceptor. Based on fluorescence quenching of IBA folding, a potassium-sensing aptasensor in the range of 0-1.4 mM was proposed. Since the quenching process was predominantly static, the binding constant and the number of binding sites were determined. In this research, based on the experimental data, the initial model of IBA G-quadruplex was constructed by molecular modeling method. The modeling structure of IBA is an intramolecular parallel-strand quadruplex conformation with two guanine tetrads. The extended molecular dynamics simulation for the model indicated that the G-quadruplex maintains its structure very well in aqueous solution in presence of K(+) in the central cavity. In contrast, it was demonstrated that the G-quadruplex structure of model in the water collapses in absence of this cation.
富含鸟嘌呤的序列在特定金属离子存在的情况下可形成G-四链体结构。在此，圆二色性、紫外可见吸收、荧光和分子动力学模拟研究表明，胰岛素结合适体（IBA）在结合K⁺后可形成分子内G-四链体结构。圆二色性（CD）光谱表明，IBA可折叠成平行G-四链体，在263nm处有一个强正峰。平衡滴定数据分析表明，阳离子结合具有协同性，K⁺的希尔系数为2.01，Na⁺的希尔系数为1.90。热变性分析表明，K⁺诱导的G-四链体比Na⁺诱导的结构更稳定。IBA折叠成G-四链体导致接触猝灭，这是由于供体和受体之间形成了非荧光复合物。基于IBA折叠的荧光猝灭，提出了一种0 - 1.4mM范围内的钾离子传感适体传感器。由于猝灭过程主要是静态的，因此测定了结合常数和结合位点的数量。在本研究中，基于实验数据，通过分子建模方法构建了IBA G-四链体的初始模型。IBA的建模结构是具有两个鸟嘌呤四联的分子内平行链四链体构象。对该模型进行的扩展分子动力学模拟表明，在中心腔存在K⁺的情况下，G-四链体在水溶液中能很好地保持其结构。相比之下，结果表明在没有这种阳离子的情况下，模型在水中的G-四链体结构会崩塌。
