Structural changes of tRNA and 5S rRNA induced with magnesium and visualized with synchrotron mediated hydroxyl radical cleavage (original) (raw)

Abstract

The structure of native yeast tRNAPhe and wheat germ ribosomal 5S RNA induced by different magnesium ion concentrations was studied in solution with a synchrotron mediated hydroxyl radical RNA cleavage reaction. We showed that very small amounts of Mg+2 can induce significant changes in the hydroxyl radical cleavage pattern of tRNAPhe. It also turned out that a reactivity of tRNAPhe towards •OH coincides with the strong metal binding sites. Because of the Mg ions are heavily hydrated one can suggest the strong correlation of the observed nucleosides reactivity in vicinity of Mg2+ binding sites with availability of water molecules as a source of hydroxyl radical. On the other hand the structure of wheat germ 5S rRNA is less sensitive to the hydroxyl radical reaction than tRNAPhe although some changes are visible at 4 mM Mg ions. It is probably due to the lack of strong Mg+2 binding sites in that molecule. The reactivity of nucleotides in loops C and D of 5S rRNA is not effected, what suggests their flexibility or involvement in higher order structure formation. There is different effect of magnesium on tRNA and 5S rRNA folding. We found that nucleotides forming strong binding sites for magnesium are very sensitive to X-ray generated hydroxyl radical and can be mapped with •OH. The results show, that guanine nucleotides are preferentially hydrated. X-ray footprinting mediated hydroxyl radical RNA cleavage is a very powerful method and has been applied to studies of stable RNAs for the first time.

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Authors and Affiliations

  1. Institute of Bioorganic Chemistry, Noskowskiego 12, 61704, Poznan, Poland
    Miroslawa Z. Barciszewska & Jan Barciszewski
  2. DESY, Notkestrasse 85, 22603, Hamburg, Germany
    Gert Rapp
  3. Arbeitsgruppe Makromolekulare Strukturanalyse, c/o DESY, Institut für Physiologische Chemie, Notkestrasse 85, 22603, Hamburg, Germany
    Christian Betzel
  4. Institut für Biochemie, Freie Universität Berlin, Thielallee 63, 14195, Berlin, Germany
    Volker A. Erdmann

Authors

  1. Miroslawa Z. Barciszewska
  2. Gert Rapp
  3. Christian Betzel
  4. Volker A. Erdmann
  5. Jan Barciszewski

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Correspondence toJan Barciszewski.

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Barciszewska, M.Z., Rapp, G., Betzel, C. et al. Structural changes of tRNA and 5S rRNA induced with magnesium and visualized with synchrotron mediated hydroxyl radical cleavage.Mol Biol Rep 28, 103–110 (2001). https://doi.org/10.1023/A:1017951120531

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