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. 1984 Nov 12;12(21):8181–8191. doi: 10.1093/nar/12.21.8181

Crosslinking transfer RNA and messenger RNA at the ribosomal decoding region: identification of the site of reaction on the messenger RNA.

G Steiner, R Lührmann, E Kuechler
PMCID: PMC320296  PMID: 6390340

Abstract

Wybutine (Ywye), situated next to the 3'-side of the anticodon of tRNAPhe from Saccharomyces cerevisiae, can be photo-crosslinked to mRNA when bound to Escherichia coli ribosomes. Crosslinking can be obtained with poly(U) as well as with oligonucleotides such as pAUGUUU or p(U)6. In order to identify the site of reaction on the mRNA, 5'-[32P]-labelled pAUGUUU was crosslinked by irradiation at 320 nm with Phe-tRNAPhe from yeast bound to the acceptor-site. The photoproduct was subsequently digested with P1-nuclease and analyzed by electrophoresis followed by homochromatography in the second dimension. As a result of the photoreaction the wybutine was found to be crosslinked to the U at the 5'-position of the corresponding UUU-codon.

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Selected References

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

  1. Baltzinger M., Fasiolo F., Remy P. Yeast phenylalanyl-tRNA synthetase. Affinity and photoaffinity labelling of the stereospecific binding sites. Eur J Biochem. 1979 Jul;97(2):481–494. doi: 10.1111/j.1432-1033.1979.tb13136.x. [DOI] [PubMed] [Google Scholar]
  2. Bertram S., Göringer U., Wagner R. Structural investigation of Phe-tRNAPhe from E.coli bound to the ribosomal A-site. Nucleic Acids Res. 1983 Feb 11;11(3):575–589. doi: 10.1093/nar/11.3.575. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Donis-Keller H., Maxam A. M., Gilbert W. Mapping adenines, guanines, and pyrimidines in RNA. Nucleic Acids Res. 1977 Aug;4(8):2527–2538. doi: 10.1093/nar/4.8.2527. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Douthwaite S., Garrett R. A., Wagner R. Comparison of Escherichia coli tRNAPhe in the free state, in the ternary complex and in the ribosomal A and P sites by chemical probing. Eur J Biochem. 1983 Mar 15;131(2):261–269. doi: 10.1111/j.1432-1033.1983.tb07258.x. [DOI] [PubMed] [Google Scholar]
  5. Jelenc P. C. Rapid purification of highly active ribosomes from Escherichia coli. Anal Biochem. 1980 Jul 1;105(2):369–374. doi: 10.1016/0003-2697(80)90472-8. [DOI] [PubMed] [Google Scholar]
  6. Kim S. H., Suddath F. L., Quigley G. J., McPherson A., Sussman J. L., Wang A. H., Seeman N. C., Rich A. Three-dimensional tertiary structure of yeast phenylalanine transfer RNA. Science. 1974 Aug 2;185(4149):435–440. doi: 10.1126/science.185.4149.435. [DOI] [PubMed] [Google Scholar]
  7. Lührmann R., Eckhardt H., Stöffler G. Codon-anticodon interaction at the ribosomal peptidyl-site. Nature. 1979 Aug 2;280(5721):423–425. doi: 10.1038/280423a0. [DOI] [PubMed] [Google Scholar]
  8. Matzke A. J., Barta A., Kuechler E. Mechanism of translocation: relative arrangement of tRNA and mRNA on the ribosome. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5110–5114. doi: 10.1073/pnas.77.9.5110. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Matzke A. J., Barta A., Kuechler E. Photo-induced crosslinking between phenylalanine transfer RNA and messenger RNA on the Escherichia coli ribosome. Eur J Biochem. 1980 Nov;112(1):169–178. doi: 10.1111/j.1432-1033.1980.tb04998.x. [DOI] [PubMed] [Google Scholar]
  10. Maxam A. M., Gilbert W. A new method for sequencing DNA. Proc Natl Acad Sci U S A. 1977 Feb;74(2):560–564. doi: 10.1073/pnas.74.2.560. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Noll M., Hapke B., Schreier M. H., Noll H. Structural dynamics of bacterial ribosomes. I. Characterization of vacant couples and their relation to complexed ribosomes. J Mol Biol. 1973 Apr 5;75(2):281–294. doi: 10.1016/0022-2836(73)90021-1. [DOI] [PubMed] [Google Scholar]
  12. Odom O. W., Craig B. B., Hardesty B. A. The conformation of the anticodon loop of yeast tRNAPhe in solution and on ribosomes. Biopolymers. 1978 Dec;17(12):2909–2931. doi: 10.1002/bip.1978.360171212. [DOI] [PubMed] [Google Scholar]
  13. Paszyc S., Rafalska M. Photochemical properties of Yt base in aqueous solution. Nucleic Acids Res. 1979 Jan;6(1):385–397. doi: 10.1093/nar/6.1.385. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Paulsen H., Robertson J. M., Wintermeyer W. Effect of ribosome binding and translocation on the anticodon of tRNAPhe as studied by wybutine fluorescence. Nucleic Acids Res. 1982 Apr 24;10(8):2651–2663. doi: 10.1093/nar/10.8.2651. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Paulsen H., Robertson J. M., Wintermeyer W. Topological arrangement of two transfer RNAs on the ribosome. Fluorescence energy transfer measurements between A and P site-bound tRNAphe. J Mol Biol. 1983 Jun 25;167(2):411–426. doi: 10.1016/s0022-2836(83)80342-8. [DOI] [PubMed] [Google Scholar]
  16. Rigler R., Wintermeyer W. Dynamics of tRNA. Annu Rev Biophys Bioeng. 1983;12:475–505. doi: 10.1146/annurev.bb.12.060183.002355. [DOI] [PubMed] [Google Scholar]
  17. Schevitz R. W., Podjarny A. D., Krishnamachari N., Hughes J. J., Sigler P. B., Sussman J. L. Crystal structure of a eukaryotic initiator tRNA. Nature. 1979 Mar 8;278(5700):188–190. doi: 10.1038/278188a0. [DOI] [PubMed] [Google Scholar]
  18. Silberklang M., Gillum A. M., RajBhandary U. L. The use of nuclease P1 in sequence analysis of end group labeled RNA. Nucleic Acids Res. 1977 Dec;4(12):4091–4108. doi: 10.1093/nar/4.12.4091. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Woese C. Molecular mechanics of translation: a reciprocating ratchet mechanism. Nature. 1970 May 30;226(5248):817–820. doi: 10.1038/226817a0. [DOI] [PubMed] [Google Scholar]
  20. Woo N. H., Roe B. A., Rich A. Three-dimensional structure of Escherichia coli initiator tRNAfMet. Nature. 1980 Jul 24;286(5771):346–351. doi: 10.1038/286346a0. [DOI] [PubMed] [Google Scholar]
  21. Wrede P., Woo N. H., Rich A. Initiator tRNAs have a unique anticodon loop conformation. Proc Natl Acad Sci U S A. 1979 Jul;76(7):3289–3293. doi: 10.1073/pnas.76.7.3289. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Wurmbach P., Nierhaus K. H. Codon-anticodon interaction at the ribosomal P (peptidyl-tRNA)site. Proc Natl Acad Sci U S A. 1979 May;76(5):2143–2147. doi: 10.1073/pnas.76.5.2143. [DOI] [PMC free article] [PubMed] [Google Scholar]

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