A rapid and sensitive assay for the detection of eukaryotic ribosome dissociation factors

David W. Russell, Linda L. Spremulli

Research output: Contribution to journalArticle

2 Scopus citations

Abstract

A rapid and sensitive assay has been developed for the factor-dependent dissociation of eukaryotic ribosomes. This assay takes advantage of the observation that initiation factor eIF-2 will bind Met-tRNAf met to 40 S subunits but not to 80 S ribosomes. Incubation of wheat germ ribosomes at 1 mm Mg2+ results in their dissociation into 40 S subunits. These subunits spontaneously reassociate when the Mg2+ concentration is raised to 4 mm. However, if the incubation at 1 mm Mg2+ is carried out in the presence of an extract containing a ribosome dissociation factor, a certain portion of the subunits will fail to reassociate when the Mg2+ concentration is raised to 4 mm. The 40 S subunits remaining due to the presence of the dissociation factor can bind [35S]Met-tRNAf met in the presence of wheat germ eIF-2. The [35S]Met-tRNAf met bound to the 40 S subunits is readily detected by its retention on a Millipore filter.

Original languageEnglish (US)
Pages (from-to)238-243
Number of pages6
JournalAnalytical biochemistry
Volume93
Issue numberC
DOIs
StatePublished - 1979

ASJC Scopus subject areas

  • Biophysics
  • Biochemistry
  • Molecular Biology
  • Cell Biology

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