title: Functional interaction between bases C1049 in domain II and G2751 in domain VI of 23S rRNA in Escherichia coli ribosomes.
abstract: The factor-binding center within the Escherichia coli ribosome is comprised of two discrete domains of 23S rRNA: the GTPase-associated region (GAR) in domain II and the sarcin-ricin loop in domain VI. These two regions appear to collaborate in the factor-dependent events that occur during protein synthesis. Current X-ray crystallography of the ribosome shows an interaction between C1049 in the GAR and G2751 in domain VI. We have confirmed this interaction by site-directed mutagenesis and chemical probing. Disruption of this base pair affected not only the chemical modification of some bases in domains II and VI and in helix H89 of domain V, but also ribosome function dependent on both EF-G and EF-Tu. Mutant ribosomes carrying the C1049 to G substitution, which show enhancement of chemical modification at G2751, were used to probe the interactions between the regions around 1049 and 2751. Binding of EF-G-GDP-fusidic acid, but not EF-G-GMP-PNP, to the ribosome protected G2751 from modification. The G2751 protection was also observed after tRNA binding to the ribosomal P and E sites. The results suggest that the interactions between the bases around 1049 and 2751 alter during different stages of the translation process.
authors.affiliation: Department of Biology, Faculty of Science, Niigata University, Niigata 950-2181, Japan.
authors.forename: Tomohiro
authors.initials: T
authors.lastname: Miyoshi
authors.forename: Toshio
authors.initials: T
authors.lastname: Uchiumi
mesh_terms: D020029:Base Pairing; D001483:Base Sequence; D004926:Escherichia coli / Q000235:genetics*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009690:Nucleic Acid Conformation; D020653:Peptide Elongation Factor G / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000737:chemistry* / Q000378:metabolism; D012343:RNA, Transfer / Q000378:metabolism; D054681:Ribosome Subunits, Large, Bacterial / Q000737:chemistry*
chemical_list: D020653:Peptide Elongation Factor G; D012338:RNA, Ribosomal, 23S; D012343:RNA, Transfer