Monitoring the structure of Escherichia coli RNase P RNA in the presence of various divalent metal ions.
Lead(II)-induced cleavage can be used as a tool to probe conformational changes in RNA. In this report, we have investigated the conformation of M1 RNA, the catalytic subunit of Escherichia coli RNase P, by studying the lead(II)-induced cleavage pattern in the presence of various divalent metal ions. Our data suggest that the overall conformation of M1 RNA is very similar in the presence of Mg(2+), Mn(2+), Ca(2+), Sr(2+) and Ba(2+), while it is changed compared to the Mg(2+)-induced conformation in the presence of other divalent metal ions, Cd(2+) for example. We also observed that correct folding of some M1 RNA domains is promoted by Pb(2+), while folding of other domain(s) requires the additional presence of other divalent metal ions, cobalt(III) hexamine or spermidine. Based on the suppression of Pb(2+) cleavage at increasing concentrations of various divalent metal ions, our findings suggest that different divalent metal ions bind with different affinities to M1 RNA as well as to an RNase P hairpin-loop substrate and yeast tRNA(Phe). We suggest that this approach can be used to obtain information about the relative binding strength for different divalent metal ions to RNA in general, as well as to specific RNA divalent metal ion binding sites. Of those studied in this report, Mn(2+) is generally among the strongest RNA binders.
Department of Cell and Molecular Biology, Box 596, Biomedical Centre, SE-751 24 Uppsala, Sweden.
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D001464:Barium / Q000494:pharmacology; D001483:Base Sequence; D002118:Calcium / Q000494:pharmacology; D020134:Catalytic Domain; D002413:Cations, Divalent / Q000494:pharmacology*; D004305:Dose-Response Relationship, Drug; D004722:Endoribonucleases / Q000235:genetics*; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D007854:Lead / Q000494:pharmacology; D008274:Magnesium / Q000494:pharmacology; D008345:Manganese / Q000494:pharmacology; D008969:Molecular Sequence Data; D009690:Nucleic Acid Conformation / Q000187:drug effects*; D012313:RNA / Q000737:chemistry* / Q000235:genetics; D016337:RNA, Catalytic / Q000235:genetics*; D043262:Ribonuclease P; D013324:Strontium / Q000494:pharmacology; D015032:Zinc / Q000494:pharmacology
D002413:Cations, Divalent; D029968:Escherichia coli Proteins; D016337:RNA, Catalytic; D001464:Barium; D007854:Lead; D008345:Manganese; D012313:RNA; D004722:Endoribonucleases; D043262:Ribonuclease P; C470865:ribonuclease P, E coli; D008274:Magnesium; D015032:Zinc; D002118:Calcium; D013324:Strontium