title: Purification, identification, and functional analysis of polysomes from the human pathogen Staphylococcus aureus.
abstract: Polysomes are macromolecular complexes made up of multiple ribosomes simultaneously translating a single mRNA into polypeptide chains. Together, the cellular mRNAs translated in this way are referred to 'translatome.' Translation determines a cell's overall gene expression profile. Studying translatome leads to a better understanding of the translational machinery and of its complex regulatory pathways. Given its fundamental role in cell homeostasis and division, bacterial translation is an important target for antibiotics. However, there are no detailed protocols for polysome purification from Staphylococcus aureus, the human pathogen responsible for the majority of multi-drug resistance issues. We therefore developed methods for the isolation of active polysomes, ribosomes, and ribosomal subunits, examining the purity and quality of each fraction and monitoring polysomal activity during protein synthesis. These steps are mandatory for the use of purified S. aureus polysomes and ribosomes for structural studies or for genome-scale analysis of most translated mRNAs.
authors.affiliation: Inserm U835 Pharmaceutical Biochemistry, University of Rennes 1, 2 avenue du Prof. Léon Bernard, 35043 Rennes, France.
authors.forename: Régine
authors.initials: R
authors.lastname: Brielle
authors.affiliation: Inserm U835 Pharmaceutical Biochemistry, University of Rennes 1, 2 avenue du Prof. Léon Bernard, 35043 Rennes, France. Electronic address: marie-laure.pinel@univ-rennes1.fr.
authors.forename: Marie-Laure
authors.initials: ML
authors.lastname: Pinel-Marie
authors.affiliation: UMR CNRS 6290 IGDR Team Translation and Folding, University of Rennes 1, Campus de Beaulieu, 35042 Rennes Cedex, France.
authors.forename: Sophie
authors.initials: S
authors.lastname: Chat
authors.affiliation: UMR CNRS 6290 IGDR Team Translation and Folding, University of Rennes 1, Campus de Beaulieu, 35042 Rennes Cedex, France; Institut Universitaire de France, France.
authors.forename: Reynald
authors.initials: R
authors.lastname: Gillet
authors.affiliation: Inserm U835 Pharmaceutical Biochemistry, University of Rennes 1, 2 avenue du Prof. Léon Bernard, 35043 Rennes, France. Electronic address: bfelden@univ-rennes1.fr.
authors.forename: Brice
authors.initials: B
authors.lastname: Felden
mesh_terms: D002458:Cell Fractionation / Q000379:methods*; D004587:Electrophoresis, Agar Gel; D008854:Microscopy, Electron; D011132:Polyribosomes / Q000737:chemistry* / Q000648:ultrastructure; D014176:Protein Biosynthesis; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D054681:Ribosome Subunits, Large, Bacterial / Q000737:chemistry* / Q000648:ultrastructure; D054680:Ribosome Subunits, Small, Bacterial / Q000737:chemistry* / Q000648:ultrastructure; D013211:Staphylococcus aureus / Q000235:genetics* / Q000378:metabolism
chemical_list: D012333:RNA, Messenger
keywords: Polysome; Ribosome; Staphylococcus aureus; Sucrose gradient purification; Translation; Translatome