title: Kinetics and mechanism of conformational changes in a G-quadruplex of thrombin-binding aptamer induced by Pb2+.
abstract: It has been shown that guanine-rich DNA can fold into a G-quadruplex with certain metal cations. The spectral characteristics, thermostability, and kinetics for the formation of a Pb(2+)-driven G-quadruplex of thrombin-binding aptamer (TBA) were measured in the current work using a combination of ultraviolet (UV) and circular dichroism (CD) spectroscopy along with stopped-flow technique. CD spectra demonstrated that TBA could fold into a unique G-quadruplex with a strong positive peak at 312 nm. Analysis of the titration data reveals that the binding stoichiometry is 1:1 for the titration of TBA with Pb(NO(3))(2), which is in accordance with the localization of the Pb(2+) ion between the adjacent G-quartets. Thermal denaturation profiles indicate that the Pb(2+)-induced intramolecular G-quadruplex is more stable than those driven by Na(+) or K(+) ions. Kinetic studies suggest that the Pb(2+)-induced folding G-quadruplex of TBA probably proceeds through the rapid formation of an intermediate Pb(2+)-TBA complex, which then isomerizes to the fully folded structure. Conformational changes transpire after the addition of Pb(NO(3))(2) to the Na(+)- or K(+)-induced G-quadruplexes, which may be attributed to the replacement of Na(+) or K(+) ions by Pb(2+) ions and the generation of a more compact structure of the Pb(2+)-TBA structure. The relaxation time, τ, of folding the G-quadruplex is reduced from 1.05 s in the presence of Pb(2+) ions alone to 0.34 s under the cooperation of initially added Na(+) ions, while τ is increased to 8.33 s under the competition of initially added K(+) ions.
authors.affiliation: Hefei National Laboratory for Physics Sciences at the Microscale, University of Science and Technology of China, Hefei, Anhui, P R China.
authors.forename: Wei
authors.initials: W
authors.lastname: Liu
authors.forename: Yan
authors.initials: Y
authors.lastname: Fu
authors.forename: Bin
authors.initials: B
authors.lastname: Zheng
authors.forename: Sheng
authors.initials: S
authors.lastname: Cheng
authors.forename: Wei
authors.initials: W
authors.lastname: Li
authors.forename: Tai-Chu
authors.initials: TC
authors.lastname: Lau
authors.forename: Haojun
authors.initials: H
authors.lastname: Liang
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000378:metabolism; D002412:Cations / Q000378:metabolism; D002942:Circular Dichroism; D054856:G-Quadruplexes; D006147:Guanine / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D007854:Lead / Q000378:metabolism*; D009691:Nucleic Acid Denaturation
chemical_list: D052157:Aptamers, Nucleotide; D002412:Cations; C081676:thrombin aptamer; D007854:Lead; D006147:Guanine