title: The RNA-mediated, asymmetric ring regulatory mechanism of the transcription termination Rho helicase decrypted by time-resolved nucleotide analog interference probing (trNAIP).
abstract: Rho is a ring-shaped, ATP-dependent RNA helicase/translocase that dissociates transcriptional complexes in bacteria. How RNA recognition is coupled to ATP hydrolysis and translocation in Rho is unclear. Here, we develop and use a new combinatorial approach, called time-resolved Nucleotide Analog Interference Probing (trNAIP), to unmask RNA molecular determinants of catalytic Rho function. We identify a regulatory step in the translocation cycle involving recruitment of the 2'-hydroxyl group of the incoming 3'-RNA nucleotide by a Rho subunit. We propose that this step arises from the intrinsic weakness of one of the subunit interfaces caused by asymmetric, split-ring arrangement of primary RNA tethers around the Rho hexamer. Translocation is at highest stake every seventh nucleotide when the weak interface engages the incoming 3'-RNA nucleotide or breaks, depending on RNA threading constraints in the Rho pore. This substrate-governed, 'test to run' iterative mechanism offers a new perspective on how a ring-translocase may function or be regulated. It also illustrates the interest and versatility of the new trNAIP methodology to unveil the molecular mechanisms of complex RNA-based systems.
authors.affiliation: Centre de Biophysique Moléculaire, CNRS UPR4301, rue Charles Sadron, 45071 Orléans cedex 2, France.
authors.forename: Emilie
authors.initials: E
authors.lastname: Soares
authors.affiliation: Centre de Biophysique Moléculaire, CNRS UPR4301, rue Charles Sadron, 45071 Orléans cedex 2, France.
authors.forename: Annie
authors.initials: A
authors.lastname: Schwartz
authors.affiliation: CNRS UMR5048, Universités Montpellier I et II, Centre de Biochimie Structurale, 29 rue de Navacelles, 34090 Montpellier, France INSERM U1054, Montpellier, France.
authors.forename: Marcello
authors.initials: M
authors.lastname: Nollmann
authors.affiliation: CNRS UMR5048, Universités Montpellier I et II, Centre de Biochimie Structurale, 29 rue de Navacelles, 34090 Montpellier, France INSERM U1054, Montpellier, France.
authors.forename: Emmanuel
authors.initials: E
authors.lastname: Margeat
authors.affiliation: Centre de Biophysique Moléculaire, CNRS UPR4301, rue Charles Sadron, 45071 Orléans cedex 2, France ITP Sciences Biologiques & Chimie du Vivant, Université d'Orléans, France marc.boudvillain@cnrs-orleans.fr.
authors.forename: Marc
authors.initials: M
authors.lastname: Boudvillain
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism*; D004247:DNA / Q000737:chemistry / Q000378:metabolism; D015336:Molecular Probe Techniques; D012313:RNA / Q000737:chemistry / Q000378:metabolism; D020365:RNA Helicases / Q000737:chemistry / Q000378:metabolism; D012234:Rho Factor / Q000737:chemistry / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D012234:Rho Factor; D012313:RNA; D004247:DNA; D020365:RNA Helicases