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. 1989 Feb 25;17(4):1475–1491. doi: 10.1093/nar/17.4.1475

New RNA-protein crosslinks in domains 1 and 2 of E. coli 30S ribosomal subunits obtained by means of an intrinsic photoaffinity probe.

E Hajnsdorf 1, A Favre 1, A Expert-Bezançon 1
PMCID: PMC331816  PMID: 2646595

Abstract

Functionally active 70S ribosomes containing 4-thiouridine (s4U) in place of uridine were prepared by a formerly described in vivo substitution method. Proteins were crosslinked to RNA by 366 nm photoactivation of s4U. We observe the systematic and characteristic formation of 30S dimers; they were eliminated for analysis of RNA-protein crosslinks. M13 probes containing rDNA inserts complementary to domains 1 and 2 of 16S RNA from the 5'end up to nucleotide 868 were used to select contiguous or overlapping RNA sections. The proteins covalently crosslinked to each RNA section were identified as S3, S4, S5, S7, S9, S18, S20 and S21. Several crosslinks are compatible with previously published sites for proteins S5, S18, S20 and S21; others for proteins S3, S4, S7, S9, S18 correspond necessarily to new sites.

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Selected References

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  1. Brewer L. A., Noller H. F. Ribonucleic acid-protein cross-linking within the intact Escherichia coli ribosome, utilizing ethylene glycol bis[3-(2-ketobutyraldehyde) ether], a reversible, bifunctional reagent: identification of 30S proteins. Biochemistry. 1983 Aug 30;22(18):4310–4315. doi: 10.1021/bi00287a023. [DOI] [PubMed] [Google Scholar]
  2. Brimacombe R., Atmadja J., Stiege W., Schüler D. A detailed model of the three-dimensional structure of Escherichia coli 16 S ribosomal RNA in situ in the 30 S subunit. J Mol Biol. 1988 Jan 5;199(1):115–136. doi: 10.1016/0022-2836(88)90383-x. [DOI] [PubMed] [Google Scholar]
  3. Caldeira de Araujo A., Favre A. Near ultraviolet DNA damage induces the SOS responses in Escherichia coli. EMBO J. 1986 Jan;5(1):175–179. doi: 10.1002/j.1460-2075.1986.tb04193.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Capel M. S., Kjeldgaard M., Engelman D. M., Moore P. B. Positions of S2, S13, S16, S17, S19 and S21 in the 30 S ribosomal subunit of Escherichia coli. J Mol Biol. 1988 Mar 5;200(1):65–87. doi: 10.1016/0022-2836(88)90334-8. [DOI] [PubMed] [Google Scholar]
  5. Chiaruttini C., Milet M., Hayes D. H., Expert-Bezancon A. Multiple crosslinks of proteins S7 and S9 to domains 3 and 4 of 16S ribosomal RNA in the Escherichia coli 30S particle. Eur J Biochem. 1986 Oct 15;160(2):363–370. doi: 10.1111/j.1432-1033.1986.tb09980.x. [DOI] [PubMed] [Google Scholar]
  6. Expert-Bezançon A., Milet M., Carbon P. Precise localization of several covalent RNA-RNA cross-link in Escherichia coli 16S RNA. Eur J Biochem. 1983 Nov 2;136(2):267–274. doi: 10.1111/j.1432-1033.1983.tb07737.x. [DOI] [PubMed] [Google Scholar]
  7. Expert-Bezançon A., Milet M., Carbon P. Precise localization of several covalent RNA-RNA cross-link in Escherichia coli 16S RNA. Eur J Biochem. 1983 Nov 2;136(2):267–274. doi: 10.1111/j.1432-1033.1983.tb07737.x. [DOI] [PubMed] [Google Scholar]
  8. GREENWOOD F. C., HUNTER W. M., GLOVER J. S. THE PREPARATION OF I-131-LABELLED HUMAN GROWTH HORMONE OF HIGH SPECIFIC RADIOACTIVITY. Biochem J. 1963 Oct;89:114–123. doi: 10.1042/bj0890114. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Hajnsdorf E., Dubreuil Y. L., Bezerra R., Favre A., Expert-Bezançon A. RNA protein crosslinks introduced into E. coli ribosomes by use of the intrinsic probe 4-thiouridine. Photochem Photobiol. 1987 Apr;45(4):445–451. doi: 10.1111/j.1751-1097.1987.tb05401.x. [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. Hayes D. H., Hayes F., Guérin M. F. Association of rapidly labelled bacterial RNA with ribosomal RNA in solutions of high ionic strength. II. J Mol Biol. 1966 Jul;18(3):499–515. doi: 10.1016/s0022-2836(66)80039-6. [DOI] [PubMed] [Google Scholar]
  12. Laemmli U. K. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. doi: 10.1038/227680a0. [DOI] [PubMed] [Google Scholar]
  13. Messing J. New M13 vectors for cloning. Methods Enzymol. 1983;101:20–78. doi: 10.1016/0076-6879(83)01005-8. [DOI] [PubMed] [Google Scholar]
  14. Millon R., Ebel J. P., Le Goffic F., Ehresmann B. Ribonucleic acid-protein crosslinking in Escherichia coli ribosomal 30S subunits by the use of two new heterobifunctional reagents: 4-azido-2,3,5,6-tetrafluoropyridine and 4-azido-3,5-dichloro-2,6-difluoropyridine. Biochem Biophys Res Commun. 1981 Aug 14;101(3):784–791. doi: 10.1016/0006-291x(81)91819-2. [DOI] [PubMed] [Google Scholar]
  15. Millon R., Olomucki M., Le Gall J. Y., Golinska B., Ebel J. P., Ehresmann B. Synthesis of a new reagent, ethyl 4-azidobenzoylaminoacetimidate, and its use for RNA-protein cross-linking within Escherichia coli ribosomal 30-S subunits. Eur J Biochem. 1980 Sep;110(2):485–492. doi: 10.1111/j.1432-1033.1980.tb04890.x. [DOI] [PubMed] [Google Scholar]
  16. Voynow P., Kurland C. G. Stoichiometry of the 30S ribosomal proteins of Escherichia coli. Biochemistry. 1971 Feb 2;10(3):517–524. doi: 10.1021/bi00779a026. [DOI] [PubMed] [Google Scholar]

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