Structure of 5 S ribosomal RNA from Escherichia coli: Identification of kethoxal-reactive sites in the A and B conformations

  • Noller H
  • Garrett R
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Abstract

The topographies of the A and B conformers of free 5 S RNA have been examined using kethoxal as a probe of single-stranded, accessible guanine residues. Each of the kethoxal-reactive guanines has been identified using diagonal electrophoresis, and the relative rate of modification at each site has been studied. Free 5 S RNA in the A form has several reactive guanines in addition to G13and G41, which are the only two available for reaction in the intact 50 S ribosomal subunit (Noller & Herr, 1974). The relative reactivities of these sites are G41≫ G13> G69> G24> G86> G107> G16, G23, G44. Modification at G23and G44reaches maximum values of only about 0.05 mol per mol 5 S RNA, suggesting that these residues are unreactive in the major conformer of the A form population. These results are compatible with a secondary structure model based on phylogenetic sequence conservation (Fox & Woese, 1975), but imply that 12 of the 18 unpaired guanines in this model are involved in further molecular interactions. The modification pattern of the B conformer demands a different base-pairing arrangement and shows that the B form contains less structure than the A form. The relative reactivities in the B form are G13> G102> G16> G24, G44> G61, G100> G23, G51, G107> G54, G56. Several sites show plateaux at submolar modification levels, indicating the existence of some conformational heterogeneity in preparations of the B form of 5 S RNA. Heat-denatured 5 S RNA appears to contain a mixture of conformers including the A and B form. These results place limitations on certain structural and functional models for 5 S RNA. For example, G44, which has often been implicated in base-pairing with tRNA, is accessible in the B form but not in the A form. Yet the B form does not bind the 5 S RNA-specific ribosomal proteins, nor is there evidence for its existence in the ribosome. © 1979.

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Authors

  • Harry F. Noller

  • Roger A. Garrett

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