title: Minimal functional structure of Escherichia coli 4.5 S RNA required for binding to elongation factor G.
abstract: Escherichia coli cells contain abundant amounts of metabolically stable 4.5 S RNA. Consisting of 114 nucleotides, 4.5 S RNA is structurally homologous to mammalian 7 S RNA, and it plays an essential role in targeting proteins containing signal peptide to the secretory apparatus by forming an signal recognition-like particle with Ffh protein. It also binds independently to protein elongation factor G (EF-G) and functions in the translation process. This RNA contains a phylogenetically conserved RNA domain, the predicted secondary structure of which consists of a hairpin motif with two bulges. We examined the binding activity of mutants with systematic deletions to define the minimal functional interaction domain of 4.5 S RNA that interacts with EF-G. This domain consisted of 35-nucleotides extending from 36 to 70 nucleotides of mature 4.5 S RNA and contained two conserved bulges in which mutations of A47, A60, G61, C62, A63, and A67 diminished binding to EF-G, whereas those at A39, C40, C41, A42, G48, and G49 did not affect binding. These data suggested that the 10 nucleotides in 4.5 S RNA, which are conserved between 4.5 S RNA and 23 S rRNA, have a key role for EF-G binding. Based on the NMR-derived structure of mutant A47U, we further verified that substituting U at A47 causes striking structural changes and the loss of the symmetrical bulge. These results indicate the mechanism by which EF-G interacts with 4.5 S RNA and the importance of the bulge structure for EF-G binding.
authors: Institute of Biological Sciences, University of Tsukuba, Tsukuba-shi, Ibaraki 305-8572, Japan. nakamura.kouji@nifty.ne.jp
authors: K
authors: K
authors: Nakamura
authors: H
authors: H
authors: Miyamoto
authors: S
authors: S
authors: Suzuma
authors: T
authors: T
authors: Sakamoto
authors: G
authors: G
authors: Kawai
authors: K
authors: K
authors: Yamane
mesh_terms: D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics*; D008969:Molecular Sequence Data; D009154:Mutation; D019906:Nuclear Magnetic Resonance, Biomolecular; D009690:Nucleic Acid Conformation; D020653:Peptide Elongation Factor G / Q000737:chemistry / Q000378:metabolism*; D011485:Protein Binding; D011487:Protein Conformation; D012329:RNA, Bacterial / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D012335:RNA, Ribosomal / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism
chemical_list: C409968:4.5S RNA; D020653:Peptide Elongation Factor G; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D011994:Recombinant Proteins