Open AccessTranslational regulation of the L11 ribosomal protein operon of Escherichia coli: analysis of the mRNA target site using oligonucleotide-directed mutagenesis.
Author(s) -
Gail Baughman,
Masayasu Nomura
Publication year - 1984
Publication title -
proceedings of the national academy of sciences of the united states of america
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 5.011
H-Index - 771
eISSN - 1091-6490
pISSN - 0027-8424
DOI - 10.1073/pnas.81.17.5389
Subject(s) - ribosomal protein , operon , translational frameshift , biology , cistron , translational regulation , ribosomal binding site , mutagenesis , ribosomal rna , ribosome , 23s ribosomal rna , genetics , messenger rna , translation (biology) , microbiology and biotechnology , mutant , gene , rna
The L11 ribosomal protein operon in Escherichia coli consists of the genes for proteins L11 and L1 and is feedback regulated by the translational repressor L1. The mRNA target site for this repression is located close to the translation initiation site of the first L11 cistron. Several mutant plasmid molecules carrying altered nucleotide sequences in the L1 target site were constructed by site-directed in vitro mutagenesis using synthetic oligodeoxyribonucleotides. Specifically, we examined the importance of a presumptive double-stranded stem structure that is common among L1 binding sites on rRNA from a variety of organisms and in L11 mRNA. Mutational alterations that disrupt the stem structure were found to abolish translational regulation as analyzed both in vitro and in vivo. Two of the mutations were combined so that the stem structure was restored but with a different primary nucleotide sequence. This double mutant was shown to restore the original phenotype, the ability to be translationally regulated by L1. These experiments show the importance of the stem structure, but not its primary sequence, for the interaction of L1 with the mRNA and support the concept that mRNA target sites share some structural features with the corresponding ribosomal protein binding sites of rRNA.