Skip to main content
RNA logoLink to RNA
. 1998 Oct;4(10):1203–1215. doi: 10.1017/s1355838298980773

Structure and stability of variants of the sarcin-ricin loop of 28S rRNA: NMR studies of the prokaryotic SRL and a functional mutant.

K Seggerson 1, P B Moore 1
PMCID: PMC1369693  PMID: 9769095

Abstract

NMR has been used to examine the conformational properties of two variants of the sarcin-ricin loop (SRL) from eukaryotic 28S rRNA, which is essential for elongation factor interactions with the ribosome: (1) its bacterial homologue, which lacks two of the bases that flank the conserved 12-nt sequence in the middle of the SRL, but which is functionally equivalent, and (2) a functionally active variant of the eukaryotic SRL in which the bulged G within the conserved sequence is replaced by an A. The data indicate that, although the bacterial SRL is less stable than the eukaryotic SRL, its conformation is closely similar. Furthermore, even though replacement of the bulged G in the SRL with an A seriously destabilizes the center of the loop, its effect on the overall conformation of the SRL appears to be modest. In the course of this work, it was serendipitously discovered that at neutral pH, the C8 proton of the bulged G, in both PRO-SRL and E73, exchanges about 10 times faster than it does in GMP.

Full Text

The Full Text of this article is available as a PDF (795.0 KB).

Selected References

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

  1. Benevides J. M., Thomas G. J., Jr Dependence of purine 8C-H exchange on nucleic acid conformation and base-pairing geometry: a dynamic probe of DNA and RNA secondary structures. Biopolymers. 1985 Apr;24(4):667–682. doi: 10.1002/bip.360240407. [DOI] [PubMed] [Google Scholar]
  2. Brandes R., Ehrenberg A. Kinetics of the proton-deuteron exchange at position H8 of adenine and guanine in DNA. Nucleic Acids Res. 1986 Dec 9;14(23):9491–9508. doi: 10.1093/nar/14.23.9491. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Correll C. C., Freeborn B., Moore P. B., Steitz T. A. Metals, motifs, and recognition in the crystal structure of a 5S rRNA domain. Cell. 1997 Nov 28;91(5):705–712. doi: 10.1016/s0092-8674(00)80457-2. [DOI] [PubMed] [Google Scholar]
  4. Dallas A., Moore P. B. The loop E-loop D region of Escherichia coli 5S rRNA: the solution structure reveals an unusual loop that may be important for binding ribosomal proteins. Structure. 1997 Dec 15;5(12):1639–1653. doi: 10.1016/s0969-2126(97)00311-0. [DOI] [PubMed] [Google Scholar]
  5. Draper D. E., Gluick T. C. Melting studies of RNA unfolding and RNA-ligand interactions. Methods Enzymol. 1995;259:281–305. doi: 10.1016/0076-6879(95)59049-8. [DOI] [PubMed] [Google Scholar]
  6. Endo Y., Tsurugi K. The RNA N-glycosidase activity of ricin A-chain. The characteristics of the enzymatic activity of ricin A-chain with ribosomes and with rRNA. J Biol Chem. 1988 Jun 25;263(18):8735–8739. [PubMed] [Google Scholar]
  7. Endo Y., Wool I. G. The site of action of alpha-sarcin on eukaryotic ribosomes. The sequence at the alpha-sarcin cleavage site in 28 S ribosomal ribonucleic acid. J Biol Chem. 1982 Aug 10;257(15):9054–9060. [PubMed] [Google Scholar]
  8. Farber N. M., Cantor C. R. A slow tritium exchange study of the solution structure of Escherichia coli 5 S ribosomal RNA. J Mol Biol. 1981 Feb 25;146(2):223–239. doi: 10.1016/0022-2836(81)90433-2. [DOI] [PubMed] [Google Scholar]
  9. Fernandez-Puentes C., Vazquez D. Effects of some proteins that inactivate the eukaryotic ribosome. FEBS Lett. 1977;78(1):143–146. doi: 10.1016/0014-5793(77)80292-5. [DOI] [PubMed] [Google Scholar]
  10. Gamble R. C., Schoemaker J. P. Rate of tritium labeling of specific purines in relation to nucleic acid and particularly transfer RNA conformation. Biochemistry. 1976 Jun 29;15(13):2791–2799. doi: 10.1021/bi00658a014. [DOI] [PubMed] [Google Scholar]
  11. Glück A., Endo Y., Wool I. G. Ribosomal RNA identity elements for ricin A-chain recognition and catalysis. Analysis with tetraloop mutants. J Mol Biol. 1992 Jul 20;226(2):411–424. doi: 10.1016/0022-2836(92)90956-k. [DOI] [PubMed] [Google Scholar]
  12. Glück A., Wool I. G. Determination of the 28 S ribosomal RNA identity element (G4319) for alpha-sarcin and the relationship of recognition to the selection of the catalytic site. J Mol Biol. 1996 Mar 15;256(5):838–848. doi: 10.1006/jmbi.1996.0130. [DOI] [PubMed] [Google Scholar]
  13. Heus H. A., Pardi A. Structural features that give rise to the unusual stability of RNA hairpins containing GNRA loops. Science. 1991 Jul 12;253(5016):191–194. doi: 10.1126/science.1712983. [DOI] [PubMed] [Google Scholar]
  14. Lane M. J., Thomas G. J., Jr Kinetics of hydrogen-deuterium exchange in guanosine 5'-monophosphate and guanosine 3':5'-monophosphate determined by laser-Raman spectroscopy. Biochemistry. 1979 Sep 4;18(18):3839–3846. doi: 10.1021/bi00585a002. [DOI] [PubMed] [Google Scholar]
  15. Liu R., Liebman S. W. A translational fidelity mutation in the universally conserved sarcin/ricin domain of 25S yeast ribosomal RNA. RNA. 1996 Mar;2(3):254–263. [PMC free article] [PubMed] [Google Scholar]
  16. Marchant A., Hartley M. R. Mutational studies on the alpha-sarcin loop of Escherichia coli 23S ribosomal RNA. Eur J Biochem. 1994 Nov 15;226(1):141–147. doi: 10.1111/j.1432-1033.1994.tb20035.x. [DOI] [PubMed] [Google Scholar]
  17. Marky L. A., Breslauer K. J. Calculating thermodynamic data for transitions of any molecularity from equilibrium melting curves. Biopolymers. 1987 Sep;26(9):1601–1620. doi: 10.1002/bip.360260911. [DOI] [PubMed] [Google Scholar]
  18. Munishkin A., Wool I. G. The ribosome-in-pieces: binding of elongation factor EF-G to oligoribonucleotides that mimic the sarcin/ricin and thiostrepton domains of 23S ribosomal RNA. Proc Natl Acad Sci U S A. 1997 Nov 11;94(23):12280–12284. doi: 10.1073/pnas.94.23.12280. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Nierhaus K. H., Schilling-Bartetzko S., Twardowski T. The two main states of the elongating ribosome and the role of the alpha-sarcin stem-loop structure of 23S RNA. Biochimie. 1992 Apr;74(4):403–410. doi: 10.1016/0300-9084(92)90118-x. [DOI] [PubMed] [Google Scholar]
  20. O'Connor M., Dahlberg A. E. The influence of base identity and base pairing on the function of the alpha-sarcin loop of 23S rRNA. Nucleic Acids Res. 1996 Jul 15;24(14):2701–2705. doi: 10.1093/nar/24.14.2701. [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Schoemaker H. J., Gamble R. C. Comparison of isotope labeling patterns of purines in three specific transfer RNAs. Biochemistry. 1976 Jun 29;15(13):2800–2803. doi: 10.1021/bi00658a015. [DOI] [PubMed] [Google Scholar]
  22. Stallings S. C., Moore P. B. The structure of an essential splicing element: stem loop IIa from yeast U2 snRNA. Structure. 1997 Sep 15;5(9):1173–1185. doi: 10.1016/s0969-2126(97)00268-2. [DOI] [PubMed] [Google Scholar]
  23. Stein E. G., Rice L. M., Brünger A. T. Torsion-angle molecular dynamics as a new efficient tool for NMR structure calculation. J Magn Reson. 1997 Jan;124(1):154–164. doi: 10.1006/jmre.1996.1027. [DOI] [PubMed] [Google Scholar]
  24. Szewczak A. A., Moore P. B., Chang Y. L., Wool I. G. The conformation of the sarcin/ricin loop from 28S ribosomal RNA. Proc Natl Acad Sci U S A. 1993 Oct 15;90(20):9581–9585. doi: 10.1073/pnas.90.20.9581. [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Szewczak A. A., Moore P. B. The sarcin/ricin loop, a modular RNA. J Mol Biol. 1995 Mar 17;247(1):81–98. doi: 10.1006/jmbi.1994.0124. [DOI] [PubMed] [Google Scholar]
  26. Tapprich W. E., Dahlberg A. E. A single base mutation at position 2661 in E. coli 23S ribosomal RNA affects the binding of ternary complex to the ribosome. EMBO J. 1990 Aug;9(8):2649–2655. doi: 10.1002/j.1460-2075.1990.tb07447.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  27. Tomasz M., Olson J., Mercado C. M. Mechanism of the isotopic exchange of the C-8 hydrogen of purines in nucleosides and in deoxyribonucleic acid. Biochemistry. 1972 Mar 28;11(7):1235–1241. doi: 10.1021/bi00757a019. [DOI] [PubMed] [Google Scholar]
  28. Wimberly B., Varani G., Tinoco I., Jr The conformation of loop E of eukaryotic 5S ribosomal RNA. Biochemistry. 1993 Feb 2;32(4):1078–1087. doi: 10.1021/bi00055a013. [DOI] [PubMed] [Google Scholar]
  29. Wool I. G., Glück A., Endo Y. Ribotoxin recognition of ribosomal RNA and a proposal for the mechanism of translocation. Trends Biochem Sci. 1992 Jul;17(7):266–269. doi: 10.1016/0968-0004(92)90407-z. [DOI] [PubMed] [Google Scholar]

Articles from RNA are provided here courtesy of The RNA Society

RESOURCES