title: Single-molecule observation of the induction of k-turn RNA structure on binding L7Ae protein.
abstract: The k-turn is a commonly occurring structural motif that introduces a tight kink into duplex RNA. In free solution, it can exist in an extended form, or by folding into the kinked structure. Binding of proteins including the L7Ae family can induce the formation of the kinked geometry, raising the question of whether this occurs by passive selection of the kinked structure, or a more active process in which the protein manipulates the RNA structure. We have devised a single-molecule experiment whereby immobilized L7Ae protein binds Cy3-Cy5-labeled RNA from free solution. We find that all bound RNA is in the kinked geometry, with no evidence for transitions to an extended form at the millisecond timescale of the camera. Furthermore, real-time binding experiments provide no evidence for a more extended intermediate even at the earliest times, at a time resolution of 16 ms. The data support a passive conformational selection model by which the protein selects a fraction of RNA that is already in the kinked conformation, thereby drawing the equilibrium into this form.
authors.affiliation: Cancer Research UK Nucleic Acid Structure Research Group, The University of Dundee, Dundee, United Kingdom.
authors.forename: Jia
authors.initials: J
authors.lastname: Wang
authors.forename: Tomáš
authors.initials: T
authors.lastname: Fessl
authors.forename: Kersten T
authors.initials: KT
authors.lastname: Schroeder
authors.forename: Jonathan
authors.initials: J
authors.lastname: Ouellet
authors.forename: Yijin
authors.initials: Y
authors.lastname: Liu
authors.forename: Alasdair D J
authors.initials: AD
authors.lastname: Freeman
authors.forename: David M J
authors.initials: DM
authors.lastname: Lilley
mesh_terms: D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D019611:Archaeoglobus fulgidus; D001483:Base Sequence; D019614:Haloarcula marismortui; D055767:Immobilized Proteins / Q000737:chemistry / Q000378:metabolism; D059372:Nucleotide Motifs; D011485:Protein Binding; D019642:RNA, Archaeal / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*
chemical_list: D019843:Archaeal Proteins; D055767:Immobilized Proteins; D019642:RNA, Archaeal