Journal of Molecular Evolution

, Volume 22, Issue 3, pp 237–242 | Cite as

Nuclease S1 analysis of eubacterial 5S rRNA secondary structure

  • M. T. MacDonell
  • R. R. Colwell


Single-strand-specific nuclease S1 was employed as a structural probe to confirm locations of unpaired nucleotide bases in 5S rRNAs purified from prokaryotic species of rRNA superfamily I. Limited nuclease S1 digests of 3′- and 5′-end-labeled [32P]5S rRNAs were electrophoresed in parallel with reference endoribonuclease digests on thin allel with reference endoribonuclease digests on thin sequencing gels. Nuclease S1 primary hydrolysis patterns were comparable for 5S rRNAs prepared from all 11 species examined in this study. The locations of base-paired regions determined by enzymatic analysis corroborate the general features of the proposed universal five-helix model for prokaryotic 5S rRNA, although the results of this study suggest a significant difference between prokaryotic and eukaryotic 5S rRNAs in the evolution of helix IV. Furthermore, the extent of base-pairing predicted by helix IV needs to be reevaluated for eubacterial species. Clipping patterns in helices II and IV appear to be consistent with a secondary structural model that undergoes a conformational rearrangement between two (or more) structures. Primary clipping patterns in the helix II region, obtained by S1 analysis, may provide useful information concerning the tertiary structure of the 5S rRNA molecule.

Key words

5S rRNA Secondary structure Nuclease S1 RNA Molecular evolution 


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  1. Brownlee GG, Sanger F, Barrell BG (1967) Nucleotide sequence of 5S ribosomal RNA fromEscherichia coli. Nature 215:735–736PubMedGoogle Scholar
  2. Corry MJ, Payne PI, Dyer TA (1974) The nucleotide sequence of 5S rRNA from the blue-green algaAnacystis nidulans. FEBS Lett 46:63–66CrossRefPubMedGoogle Scholar
  3. Delihas N, Andresini W, Andersen J, Berns D (1982) Structural features unique to the 5S ribosomal RNAs of the thermophilic cyanobacteriumSynechococcus lividus III and the green plant chloroplasts. J Mol Biol 162:721–727CrossRefPubMedGoogle Scholar
  4. De Vos P, De Ley J (1983) Intra- and intergeneric similarities ofPseudomonas andXanthomonas ribosomal ribonucleic acid cistrons. Int J Syst Bacteriol 33:487–509Google Scholar
  5. De Wachter R, Chen M-W, Vandenberghe A (1982) Conservation of secondary structure in 5S ribosomal RNA: a uniform model for eukaryotic, eubacterial, archaebacterial and organelle sequences is energetically favourable. Biochimie 64: 311–329PubMedGoogle Scholar
  6. Digweed M, Ulbrich N, Erdmann VA (1984) The secondary structure of the 5S rRNA from the horsetailEquisetum arvense. FEBS Lett 176:255–260CrossRefGoogle Scholar
  7. Douthwaite S, Garrett RA (1981) Secondary structure of prokaryotic 5S ribosomal ribonucleic acids: a study with ribonucleases. Biochemistry 20:7301–7307CrossRefPubMedGoogle Scholar
  8. Erdmann VA, Wolters J, Huysmans E, Vandenberghe A, De Wachter R (1984) Collection of published 5S and 5.8S ribosomal RNA sequences. Nucleic Acids Res 12:r133-r166PubMedGoogle Scholar
  9. Garrett-Wheeler E, Lockart RE, Kumar A (1984) Mapping of psoralen cross-linked nucleotides in RNA. Nucleic Acids Res 7:3405–3423Google Scholar
  10. Hansen JN (1981) Use of solubilizable acrylamide disulfide gels for isolation of DNA fragments suitable for sequence analysis. Anal Biochem 116:146–151CrossRefPubMedGoogle Scholar
  11. Kao TH, Crothers DM (1980) A proton-coupled conformational switch ofEscherichia coli 5S ribosomal RNA. Proc Natl Acad Sci USA 77:3360–3364PubMedGoogle Scholar
  12. MacDonell MT, Colwell RR (1984a) The nucleotide base sequence of Vibronacease 5S rRNA. FEBS Lett 175:183–188CrossRefPubMedGoogle Scholar
  13. MacDonell MT, Colwell RR (1984b) Nucleotide sequence of 5S ribosomal RNA fromVibrio marinus. Microbiol Sci 1: 229–231PubMedGoogle Scholar
  14. MacDonell MT, Colwell, RR (1985) Phylogeny of the Vibrionaceae and recommendation for two new generaListonella andShewanella. Syst Appl Microbiol 6:171–182Google Scholar
  15. MacKay RM, Salgado D, Bonen L, Stackebrandt E, Doolittle WF (1982) The 5S ribosomal RNAs ofParacoccus denitrificans andProchloron. Nucleic Acids Res 10:2963–2970PubMedGoogle Scholar
  16. Ninio J (1979) Prediction of pairing schemes in RNA molecules—loop contributions and energy of wobble and nonwobble pairs. Biochimie 61:1133–1150PubMedGoogle Scholar
  17. Noller HF (1974) Topography of 16S RNA in 30S ribosomal subunits. Nucleotide sequences and location of sites of reaction with kethoxal. Biochemistry 13:4694–4703PubMedGoogle Scholar
  18. Noller HF (1984) Structure of ribosomal RNA. Annu Rev Biochem 53:119–162CrossRefPubMedGoogle Scholar
  19. Peattie DA, Gilbert W (1980) Chemical probes for higher-order structure in RNA. Proc. Natl Acad Sci USA 77:4679–4682PubMedGoogle Scholar
  20. Pieler T, Erdmann VA (1982) Three dimensional structural model of eubacterial 5S RNA that has functional implications. Proc Natl Acad Sci USA 79:4599–4603PubMedGoogle Scholar
  21. Pieler T, Schreiber A, Erdmann VA (1984) Comparative structural analysis of eubacterial 5S rRNA by oxidation of adenines in the N-1 position. Nucleic Acids Res 12:3115–3126PubMedGoogle Scholar
  22. Rabin D, Crothers DM (1979) Analysis of RNA secondary structure by photochemical reversal of psoralen crosslinks. Nucleic Acids Res 7:689–703PubMedGoogle Scholar
  23. Randerath K, Randerath E (1967) Thin-layer separation methods for nucleic acid derivatives. Methods Enzymol 12A:323–347Google Scholar
  24. Ross A, Brimacombe R (1979) Experimental determination of interacting sequences in ribosomal RNA. Nature 281:271–276PubMedGoogle Scholar
  25. Sanger F, Coulsen AR (1978) The use of thin acrylamide gels for sequencing. FEBS Lett 87:107–110CrossRefPubMedGoogle Scholar
  26. Trifonov EN, Bolshoi G (1983) Open and closed 5S ribosomal RNA, the only two universal structures encoded in the nucleotide sequences. J Mol Biol 169:1–13PubMedGoogle Scholar
  27. Vogt VM (1980) Purification and properties of S1 nuclease fromAspergillus. Methods Enzymol 65:248–264PubMedGoogle Scholar

Copyright information

© Springer-Verlag 1985

Authors and Affiliations

  • M. T. MacDonell
    • 1
  • R. R. Colwell
    • 1
  1. 1.Department of MicrobiologyThe University of MarylandCollege ParkUSA

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