title: A large collapsed-state RNA can exhibit simple exponential single-molecule dynamics.
abstract: The process of large RNA folding is believed to proceed from many collapsed structures to a unique functional structure requiring precise organization of nucleotides. The diversity of possible structures and stabilities of large RNAs could result in non-exponential folding kinetics (e.g. stretched exponential) under conditions where the molecules have not achieved their native state. We describe a single-molecule fluorescence resonance energy transfer (FRET) study of the collapsed-state region of the free energy landscape of the catalytic domain of RNase P RNA from Bacillus stearothermophilus (C(thermo)). Ensemble measurements have shown that this 260 residue RNA folds cooperatively to its native state at >or=1 mM Mg(2+), but little is known about the conformational dynamics at lower ionic strength. Our measurements of equilibrium conformational fluctuations reveal simple exponential kinetics that reflect a small number of discrete states instead of the expected inhomogeneous dynamics. The distribution of discrete dwell times, collected from an "ensemble" of 300 single molecules at each of a series of Mg(2+) concentrations, fit well to a double exponential, which indicates that the RNA conformational changes can be described as a four-state system. This finding is somewhat unexpected under [Mg(2+)] conditions in which this RNA does not achieve its native state. Observation of discrete well-defined conformations in this large RNA that are stable on the seconds timescale at low [Mg(2+)] (<0.1 mM) suggests that even at low ionic strength, with a tremendous number of possible (weak) interactions, a few critical interactions may produce deep energy wells that allow for rapid averaging of motions within each well, and yield kinetics that are relatively simple.
authors.affiliation: Department of Chemistry, University of Chicago, Chicago, Illinois 60637, USA.
authors.forename: Glenna J
authors.initials: GJ
authors.lastname: Smith
authors.forename: Kang Taek
authors.initials: KT
authors.lastname: Lee
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Qu
authors.forename: Zheng
authors.initials: Z
authors.lastname: Xie
authors.forename: Jelena
authors.initials: J
authors.lastname: Pesic
authors.forename: Tobin R
authors.initials: TR
authors.lastname: Sosnick
authors.forename: Tao
authors.initials: T
authors.lastname: Pan
authors.forename: Norbert F
authors.initials: NF
authors.lastname: Scherer
mesh_terms: D001483:Base Sequence; D001411:Geobacillus stearothermophilus / Q000201:enzymology; D008274:Magnesium / Q000737:chemistry; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D012313:RNA / Q000737:chemistry* / Q000378:metabolism*; D013997:Time Factors
chemical_list: D012313:RNA; D008274:Magnesium