PMID:11294875
| Citation |
Nakamura, K, Miyamoto, H, Suzuma, S, Sakamoto, T, Kawai, G and Yamane, K (2001) Minimal functional structure of Escherichia coli 4.5 S RNA required for binding to elongation factor G. J. Biol. Chem. 276:22844-9 |
|---|---|
| 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. |
| Links |
PubMed Online version:10.1074/jbc.M101376200 |
| Keywords |
Base Sequence; Escherichia coli/genetics; Molecular Sequence Data; Mutation; Nuclear Magnetic Resonance, Biomolecular; Nucleic Acid Conformation; Peptide Elongation Factor G/chemistry; Peptide Elongation Factor G/metabolism; Protein Binding; Protein Conformation; RNA, Bacterial/chemistry; RNA, Bacterial/isolation & purification; RNA, Bacterial/metabolism; RNA, Ribosomal/chemistry; RNA, Ribosomal/isolation & purification; RNA, Ribosomal/metabolism; Recombinant Proteins/chemistry; Recombinant Proteins/metabolism |
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