title: In vivo tmRNA protection by SmpB and pre-ribosome binding conformation in solution.
abstract: TmRNA is an abundant RNA in bacteria with tRNA and mRNA features. It is specialized in trans-translation, a translation rescuing system. We demonstrate that its partner protein SmpB binds the tRNA-like region (TLD) in vivo and chaperones the fold of the TLD-H2 region. We use an original approach combining the observation of tmRNA degradation pathways in a heterologous system, the analysis of the tmRNA digests by MS and NMR, and co-overproduction assays of tmRNA and SmpB. We study the conformation in solution of tmRNA alone or in complex with one SmpB before ribosome binding using SAXS. Our data show that Mg(2+) drives compaction of the RNA structure and that, in the absence of Mg(2+), SmpB has a similar effect albeit to a lesser extent. Our results show that tmRNA is intrinsically structured in solution with identical topology to that observed on complexes on ribosomes which should facilitate its subsequent recruitment by the 70S ribosome, free or preloaded with one SmpB molecule.
authors.affiliation: CNRS-UMR 8015, Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, 75270 Paris Cedex 06, France Université Paris Descartes, LCRB, Faculté de Pharmacie, 75270 Paris Cedex 06, France.
authors.forename: Ehsan
authors.initials: E
authors.lastname: Ranaei-Siadat
authors.affiliation: Université Paris-Sud, IBBMC, UMR8619, 91405 Orsay, France CNRS, 91405 Orsay, France.
authors.forename: Cécile
authors.initials: C
authors.lastname: Mérigoux
authors.affiliation: CNRS-UMR 8015, Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, 75270 Paris Cedex 06, France Université Paris Descartes, LCRB, Faculté de Pharmacie, 75270 Paris Cedex 06, France.
authors.forename: Bili
authors.initials: B
authors.lastname: Seijo
authors.affiliation: CNRS-UMR 8015, Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, 75270 Paris Cedex 06, France Université Paris Descartes, LCRB, Faculté de Pharmacie, 75270 Paris Cedex 06, France.
authors.forename: Luc
authors.initials: L
authors.lastname: Ponchon
authors.affiliation: CNRS, IPHC-LSMBO, Université Louis Pasteur Bât, 67087 Strasbourg, France.
authors.forename: Jean-Michel
authors.initials: JM
authors.lastname: Saliou
authors.affiliation: AMIB, INRIA Saclay-Île de France, 91120 Palaiseau, France LIX, CNRS UMR 7161, École Polytechnique, 91120 Palaiseau, France.
authors.forename: Julie
authors.initials: J
authors.lastname: Bernauer
authors.affiliation: CNRS, IPHC-LSMBO, Université Louis Pasteur Bât, 67087 Strasbourg, France.
authors.forename: Sarah
authors.initials: S
authors.lastname: Sanglier-Cianférani
authors.affiliation: CNRS-UMR 8015, Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, 75270 Paris Cedex 06, France Université Paris Descartes, LCRB, Faculté de Pharmacie, 75270 Paris Cedex 06, France.
authors.forename: Fréderic
authors.initials: F
authors.lastname: Dardel
authors.affiliation: Université Paris-Sud, IBBMC, UMR8619, 91405 Orsay, France CNRS, 91405 Orsay, France sylvie.nonin@parisdescartes.fr Patrice.Vachette@u-psud.fr.
authors.forename: Patrice
authors.initials: P
authors.lastname: Vachette
authors.affiliation: CNRS-UMR 8015, Laboratoire de Cristallographie et RMN Biologiques, Faculté de Pharmacie, 75270 Paris Cedex 06, France Université Paris Descartes, LCRB, Faculté de Pharmacie, 75270 Paris Cedex 06, France sylvie.nonin@parisdescartes.fr Patrice.Vachette@u-psud.fr.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Nonin-Lecomte
mesh_terms: D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D009690:Nucleic Acid Conformation; D011485:Protein Binding; D014176:Protein Biosynthesis; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000737:chemistry* / Q000378:metabolism*; D016601:RNA-Binding Proteins / Q000378:metabolism*; D012270:Ribosomes / Q000378:metabolism*; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D014961:X-Ray Diffraction
chemical_list: D012329:RNA, Bacterial; D016601:RNA-Binding Proteins; C430328:small protein B; C096222:tmRNA
keywords: NMR; SAXS; SmpB; in vivo protection; tmRNA; trans-translation