Osmolytes stimulate the reconstitution of functional 50S ribosomes from in vitro transcripts of Escherichia coli 23S rRNA

Katharina Semrad, Rachel Green

Research output: Contribution to journalArticle

Abstract

Functional Escherichia coli 50S ribosomal subunits can be reconstituted from their natural rRNA and protein components. However, when the assembly is performed with in vitro-transcribed 23S rRNA, the reconstitution efficiency is diminished by four orders of magnitude. We tested a variety of chemical chaperones (compounds that are typically used for protein folding), putative RNA chaperones (proteins) and ribosome-targeted antibiotics (small-molecule ligands) that might be reasoned to aid in folding and assembly. Addition of the osmolyte trimethylamine-oxide (TMAO) and the ketolide antibiotic telithromycin (HMR3647) to the reconstitution stimulates its efficiency up to 100-fold yielding a substantially improved system for the in vitro analysis of mutant ribosomes.

Original languageEnglish (US)
Pages (from-to)401-411
Number of pages11
JournalRNA
Volume8
Issue number4
DOIs
StatePublished - 2002

Keywords

  • In vitro reconstitution
  • RNA folding
  • Ribosome assembly
  • Telithromycin
  • Trimethylamine oxide (TMAO)

ASJC Scopus subject areas

  • Molecular Biology

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