Relation between the ribosomal sites involved in initiation and elongation of polypeptide chains. Evidence for two guanosine triphosphatase sites

A. H. Lockwood, U. Maitra

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Abstract

The relationships between ribosomal sites involved in initiation and elongation of polypeptide chains have been investigated. Thiostrepton, a peptide antibiotic that binds to a single site on the 50 S ribosomal subunit, prevents subsequent interaction of the ribosome with peptide chain elongation factors G (EF G) and Tu (EF Tu). In contrast, the drug does not directly impair binding of formyl methionyl transfer RNA to ribosomes. Bound formyl methionyl transfer RNA can react with puromycin indicating formation of a functional 70 S initiation complex. Prior binding of EF G to ribosomes in the presence of either 5' guanylyl methylene diphosphonate or fusidic acid and guanosine diphosphate prevents the subsequent binding of the ternary complex, aminoacyl transfer RNA EF Tu guanosine triphosphate to such ribosomes. In contrast, formation of the polypeptide initiation complex on ribosomes carrying bound EF G is not inhibited but is somewhat stimulated. At a low molar ratio of IF 2 to ribosomes, prior binding of either thiostrepton or EF G to ribosomes does not inhibit ribosome dependent uncoupled hydrolysis of guanosine triphosphate catalyzed by IF 2. At a high molar ratio of IF 2 to ribosomes however, the IF 2 guanosine triphosphatase activity is partially inhibited by binding of either thiostrepton or EF G. However, maximum inhibition does not exceed 50%. The inhibition observed is on the rate of guanosine triphosphate hydrolysis; the yield of the hydrolytic reaction is not impaired. These results suggest that while aminoacyl transfer RNA EF Tu guanosine triphosphate complex and EF G interact with either the same or overlapping regions of the ribosome, the ribosomal site for uncoupled hydrolysis of guanosine triphosphate catalyzed by IF 2 does not overlap the EF G site. This implies the existence of at least two guanosine triphosphatase sites on ribosomes: an IF 2 specific site and an EF G site. However, at a relatively high molar excess of IF 2 to ribosomes, IF 2 can also interact with the EF G site.

Original languageEnglish (US)
Pages (from-to)346-352
Number of pages7
JournalJournal of Biological Chemistry
Volume249
Issue number2
Publication statusPublished - Jan 1 1974

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ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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