Skip to main content
PMC home page
Nucleic Acids Research logoLink to Nucleic Acids Research
. 1987 May 11;15(9):3653–3670. doi: 10.1093/nar/15.9.3653

Protein binding sites on Escherichia coli 16S RNA; RNA regions that are protected by proteins S7, S14 and S19 in the presence or absence of protein S9.

L Wiener, R Brimacombe
PMCID: PMC340774  PMID: 2438658

Abstract

14C-labelled proteins from E. coli 30S ribosomal subunits were isolated by HPLC, and selected groups of these proteins were reconstituted with 32P-labelled 16S RNA. The isolated reconstituted particles were partially digested with ribonuclease A, and the RNA fragments protected by the proteins were separated by gel electrophoresis and subjected to sequence analysis. Protein S7 alone gave no protected fragments, but S7 together with S14 and S19 protected an RNA region comprising the sequences 936-965, 972-1030, 1208-1262 and 1285-1379 of the 16S RNA. Addition of increasing amounts of protein S9 to the S7/S14/S19 particle resulted in a parallel increase in the protection of the hairpin loop between bases 1262 and 1285. The results are discussed in terms of the three-dimensional folding of 16S RNA in the 30S subunit.

Full text

PDF
3653

Images in this article

3660Image
on p.3660
3661Image
on p.3661
3663Image
on p.3663

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Atmadja J., Brimacombe R., Blöcker H., Frank R. Investigation of the tertiary folding of Escherichia coli 16S RNA by in situ intra-RNA cross-linking within 30S ribosomal subunits. Nucleic Acids Res. 1985 Oct 11;13(19):6919–6936. doi: 10.1093/nar/13.19.6919. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Breitenreuter G., Lotti M., Stöffler-Meilicke M., Stöffler G. Comparative electron microscopic study on the location of ribosomal proteins S3 and S7 on the surface of the E. coli 30S subunit using monoclonal and conventional antibody. Mol Gen Genet. 1984;197(2):189–195. doi: 10.1007/BF00330962. [DOI] [PubMed] [Google Scholar]
  3. Brimacombe R., Maly P., Zwieb C. The structure of ribosomal RNA and its organization relative to ribosomal protein. Prog Nucleic Acid Res Mol Biol. 1983;28:1–48. doi: 10.1016/s0079-6603(08)60081-1. [DOI] [PubMed] [Google Scholar]
  4. Brockmöller J., Kamp R. M. Isolation and identification of two protein-protein crosslinks within the two subunits of Bacillus stearothermophilus ribosomes. Biol Chem Hoppe Seyler. 1986 Sep;367(9):925–935. doi: 10.1515/bchm3.1986.367.2.925. [DOI] [PubMed] [Google Scholar]
  5. Brosius J., Palmer M. L., Kennedy P. J., Noller H. F. Complete nucleotide sequence of a 16S ribosomal RNA gene from Escherichia coli. Proc Natl Acad Sci U S A. 1978 Oct;75(10):4801–4805. doi: 10.1073/pnas.75.10.4801. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Carbon P., Ehresmann C., Ehresmann B., Ebel J. P. The complete nucleotide sequence of the ribosomal 16-S RNA from Excherichia coli. Experimental details and cistron heterogeneities. Eur J Biochem. 1979 Oct 15;100(2):399–410. doi: 10.1111/j.1432-1033.1979.tb04183.x. [DOI] [PubMed] [Google Scholar]
  7. Dohme F., Nierhaus K. H. Total reconstitution and assembly of 50 S subunits from Escherichia coli Ribosomes in vitro. J Mol Biol. 1976 Nov 15;107(4):585–599. doi: 10.1016/s0022-2836(76)80085-x. [DOI] [PubMed] [Google Scholar]
  8. Geyl D., Böck A., Isono K. An improved method for two-dimensional gel-electrophoresis: analysis of mutationally altered ribosomal proteins of Escherichia coli. Mol Gen Genet. 1981;181(3):309–312. doi: 10.1007/BF00425603. [DOI] [PubMed] [Google Scholar]
  9. Gregory R. J., Zeller M. L., Thurlow D. L., Gourse R. L., Stark M. J., Dahlberg A. E., Zimmermann R. A. Interaction of ribosomal proteins S6, S8, S15 and S18 with the central domain of 16 S ribosomal RNA from Escherichia coli. J Mol Biol. 1984 Sep 15;178(2):287–302. doi: 10.1016/0022-2836(84)90145-1. [DOI] [PubMed] [Google Scholar]
  10. Hardy S. J., Kurland C. G., Voynow P., Mora G. The ribosomal proteins of Escherichia coli. I. Purification of the 30S ribosomal proteins. Biochemistry. 1969 Jul;8(7):2897–2905. doi: 10.1021/bi00835a031. [DOI] [PubMed] [Google Scholar]
  11. Held W. A., Ballou B., Mizushima S., Nomura M. Assembly mapping of 30 S ribosomal proteins from Escherichia coli. Further studies. J Biol Chem. 1974 May 25;249(10):3103–3111. [PubMed] [Google Scholar]
  12. Huber P. W., Wool I. G. Nuclease protection analysis of ribonucleoprotein complexes: use of the cytotoxic ribonuclease alpha-sarcin to determine the binding sites for Escherichia coli ribosomal proteins L5, L18, and L25 on 5S rRNA. Proc Natl Acad Sci U S A. 1984 Jan;81(2):322–326. doi: 10.1073/pnas.81.2.322. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Kaltschmidt E., Stöffler G., Dzionara M., Wittmann H. G. Ribosomal proteins. XVII. Comparative studies on ribosomal proteins of four strains of Escherichia coli. Mol Gen Genet. 1970;109(4):303–308. doi: 10.1007/BF00267700. [DOI] [PubMed] [Google Scholar]
  14. Kamp R. M., Bosserhoff A., Kamp D., Wittmann-Liebold B. Application of high-performance liquid chromatographic techniques to the separation of ribosomal proteins of different organisms. J Chromatogr. 1984 Dec 28;317:181–192. doi: 10.1016/s0021-9673(01)91658-9. [DOI] [PubMed] [Google Scholar]
  15. Kyriatsoulis A., Maly P., Greuer B., Brimacombe R., Stöffler G., Frank R., Blöcker H. RNA-protein cross-linking in Escherichia coli ribosomal subunits: localization of sites on 16S RNA which are cross-linked to proteins S17 and S21 by treatment with 2-iminothiolane. Nucleic Acids Res. 1986 Feb 11;14(3):1171–1186. doi: 10.1093/nar/14.3.1171. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Lake J. A. Ribosome evolution: the structural bases of protein synthesis in archaebacteria, eubacteria, and eukaryotes. Prog Nucleic Acid Res Mol Biol. 1983;30:163–194. doi: 10.1016/s0079-6603(08)60686-8. [DOI] [PubMed] [Google Scholar]
  17. Maly P., Brimacombe R. Refined secondary structure models for the 16S and 23S ribosomal RNA of Escherichia coli. Nucleic Acids Res. 1983 Nov 11;11(21):7263–7286. doi: 10.1093/nar/11.21.7263. [DOI] [PMC free article] [PubMed] [Google Scholar]
  18. Maly P., Rinke J., Ulmer E., Zwieb C., Brimacombe R. Precise localization of the site of cross-linking between protein L4 and 23S ribonucleic acid induced by mild ultraviolet irradiation of Escherichia coli 50S ribosomal subunits. Biochemistry. 1980 Sep 2;19(18):4179–4188. doi: 10.1021/bi00559a007. [DOI] [PubMed] [Google Scholar]
  19. Morgan J., Brimacombe R. A series of specific ribonucleoprotein fragments from the 30-S subparticle of Escherichia coli ribosomes. Eur J Biochem. 1972 Sep 25;29(3):542–552. doi: 10.1111/j.1432-1033.1972.tb02020.x. [DOI] [PubMed] [Google Scholar]
  20. Möller K., Brimacombe R. Specific cross-linking of proteins S7 and L4 to ribosomal RNA, by UV irradiation of Escherichia coli ribosomal subunits. Mol Gen Genet. 1975 Dec 9;141(4):343–355. doi: 10.1007/BF00331455. [DOI] [PubMed] [Google Scholar]
  21. Stiege W., Atmadja J., Zobawa M., Brimacombe R. Investigation of the tertiary folding of Escherichia coli ribosomal RNA by intra-RNA cross-linking in vivo. J Mol Biol. 1986 Sep 5;191(1):135–138. doi: 10.1016/0022-2836(86)90429-8. [DOI] [PubMed] [Google Scholar]
  22. Stiege W., Zwieb C., Brimacombe R. Precise localisation of three intra-RNA cross-links in 23S RNA and one in 5S RNA, induced by treatment of Escherichia coli 50S ribosomal subunits with bis-(2-chloroethyl)-methylamine. Nucleic Acids Res. 1982 Nov 25;10(22):7211–7229. doi: 10.1093/nar/10.22.7211. [DOI] [PMC free article] [PubMed] [Google Scholar]
  23. Ulmer E., Meinke M., Ross A., Fink G., Brimacombe R. Chemical cross-linking of protein to RNA within intact ribosomal subunits from Escherichia coli. Mol Gen Genet. 1978 Apr 6;160(2):183–193. doi: 10.1007/BF00267480. [DOI] [PubMed] [Google Scholar]
  24. Ungewickell E., Garrett R., Ehresmann C., Stiegler P., Fellner P. An investigation of the 16-S RNA binding sites of ribosomal proteins S4, S8, S15, and S20 FROM Escherichia coli. Eur J Biochem. 1975 Feb 3;51(1):165–180. doi: 10.1111/j.1432-1033.1975.tb03917.x. [DOI] [PubMed] [Google Scholar]
  25. Volckaert G., Fiers W. Micro thin-layer techniques for rapid sequence analysis of 32P-labeled RNA: double digestion and pancreatic ribonuclease analyses. Anal Biochem. 1977 Nov;83(1):228–239. doi: 10.1016/0003-2697(77)90531-0. [DOI] [PubMed] [Google Scholar]
  26. Wower I., Brimacombe R. The localization of multiple sites on 16S RNA which are cross-linked to proteins S7 and S8 in Escherichia coli 30S ribosomal subunits by treatment with 2-iminothiolane. Nucleic Acids Res. 1983 Mar 11;11(5):1419–1437. doi: 10.1093/nar/11.5.1419. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Yuki A., Brimacombe R. Nucleotide sequences of Escherichia coli 16-S RNA associated with ribosomal proteins S7, S9, S10, S14 and S19. Eur J Biochem. 1975 Aug 1;56(1):23–34. doi: 10.1111/j.1432-1033.1975.tb02203.x. [DOI] [PubMed] [Google Scholar]

Articles from Nucleic Acids Research are provided here courtesy of Oxford University Press

RESOURCES