Base changes at position 792 of Escherichia coli 16S rRNA affect assembly of 70S ribosomes

Proc Natl Acad Sci U S A. 1990 May;87(10):3700-4. doi: 10.1073/pnas.87.10.3700.

Abstract

To investigate the function of base 792 of 16S rRNA in 30S ribosomes of Escherichia coli, the wild-type (adenine) residue was changed to guanine, cytosine, or uracil by oligonucleotide-directed mutagenesis. Each base change conferred a unique phenotype on the cells. Cells containing plasmid pKK3535 with G792 or T792 showed no difference in generation time in LB broth containing ampicillin, whereas cells with C792 exhibited a 20% increase in generation time in this medium. To study the effect on cell growth of a homogeneous population of mutant ribosomes, the mutations were cloned into the 16S rRNA gene on pKK3535 carrying a spectinomycin-resistance marker (thymine at position 1192), and the cells were grown with spectinomycin. Cells containing G792 or C792 showed 16% and 56% increases in generation time, respectively, and a concomitant decrease in 35S assimilation into proteins. Cells with T792 did not grow in spectinomycin-containing medium. Maxicell analyses indicated decreasing ability to form 70S ribosomes from 30S subunits containing guanine, cytosine, or uracil at position 792 in 16S rRNA. It appeared that C792-containing 30S ribosomes had lost the ability to bind initiation factor 3.

Publication types

  • Research Support, U.S. Gov't, Non-P.H.S.
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • Base Sequence
  • Drug Resistance, Microbial / genetics
  • Escherichia coli / genetics*
  • Escherichia coli / growth & development
  • Escherichia coli / metabolism
  • Genes, Bacterial
  • Molecular Sequence Data
  • Mutation
  • Nucleic Acid Conformation
  • Plasmids
  • RNA, Ribosomal / genetics*
  • RNA, Ribosomal, 16S / genetics*
  • Ribosomes / metabolism*
  • Spectinomycin / pharmacology

Substances

  • RNA, Ribosomal
  • RNA, Ribosomal, 16S
  • Spectinomycin