title: Role of conserved nucleotides in building the 16S rRNA binding site of E. coli ribosomal protein S8.
abstract: Ribosomal protein S8 specifically recognizes a helical and irregular region of 16S rRNA that is highly evolutionary constrained. Despite its restricted size, the precise conformation of this region remains a question of debate. Here, we used chemical probing to analyze the structural consequences of mutations in this RNA region. These data, combined with computer modelling and previously published data on protein binding were used to investigate the conformation of the RNA binding site. The experimental data confirm the model in which adenines A595, A640 and A642 bulge out in the deep groove. In addition to the already proposed non canonical U598-U641 interaction, the structure is stabilized by stacking interactions (between A595 and A640) and an array of hydrogen bonds involving bases and the sugar phosphate backbone. Mutations that alter the ability to form these interdependent interactions result in a local destabilization or reorganization. The specificity of recognition by protein S8 is provided by the irregular and distorted backbone and the two bulged adenines 640 and 642 in the deep groove. The third adenine (A595) is not a direct recognition site but must adopt a bulged position. The U598-U641 pair should not be directly in contact with the protein.
authors.affiliation: UPR 9002 du CNRS, Institut de Biologie Moléculaire et Cellulaire, Strasbourg, France.
authors.forename: C
authors.initials: C
authors.lastname: Allmang
authors.forename: M
authors.initials: M
authors.lastname: Mougel
authors.forename: E
authors.initials: E
authors.lastname: Westhof
authors.forename: B
authors.initials: B
authors.lastname: Ehresmann
authors.forename: C
authors.initials: C
authors.lastname: Ehresmann
mesh_terms: D001483:Base Sequence; D001665:Binding Sites; D003198:Computer Simulation; D017124:Conserved Sequence; D004926:Escherichia coli / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D017354:Point Mutation / Q000502:physiology; D012336:RNA, Ribosomal, 16S / Q000737:chemistry* / Q000378:metabolism*; D012269:Ribosomal Proteins / Q000378:metabolism*
chemical_list: D012336:RNA, Ribosomal, 16S; D012269:Ribosomal Proteins; C030254:ribosomal protein S8