Functional effects of mutating the closing GxA base-pair of a conserved hairpin loop in 23 S ribosomal RNA.
Recently, Draper and co-workers solved the structure of a hexanucleotide hairpin loop that is conserved in large subunit ribosomal RNAs. (In Escherichia coli, the hexanucleotide consists of nucleotides 1093 to 1098, in the GTPase center of 23 S rRNA.) A major feature of that structure is a G1093xA1098 base-pair that closes the loop. Our laboratory reported previously the isolation of the mutation G1093A and its characterization as a suppressor of UGA mutations and a cause of temperature-conditional lethality. For the work reported here, we asked whether G1093A causes its phenotypes precisely because it is part of the G1093/A1098 base-pair. Using oligonucleotide-directed site-specific mutagenesis, we introduced base substitutions at nucleotides 1093 and 1098 into a plasmid-borne ribosomal RNA operon (rrnB). Each mutant plasmid was then tested for the two mutant phenotypes, nonsense suppression and temperature-dependent growth inhibition. Our results indicate that mutations at 1093 do not cause the mutant phenotypes because G1093 is part of the G1093xA1098 base-pair. We discuss alternative avenues to the observed mutant phenotypes and, in particular, present a model in which a specific interaction of the loop is involved in peptide chain termination.
Xu
W
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Department of Molecular Genetics, The University of Texas, Houston 77030, USA.
Murgola
E J
EJ
D001482:Base Composition; D001483:Base Sequence; D017124:Conserved Sequence; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development; D006151:Guanosine; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation; D010641:Phenotype; D017354:Point Mutation; D011132:Polyribosomes / Q000378:metabolism; D012338:RNA, Ribosomal, 23S / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D013489:Suppression, Genetic
D012338:RNA, Ribosomal, 23S; D006151:Guanosine