title: Altered lead(II)-cleavage pattern of free Phe-tRNAPhe and Phe-tRNAPhe in ternary complex with EF-Tu:GTP.
abstract: Pb2+ ions in sub-millimolar concentrations are known to cleave internucleotide bonds of phenylalanine-specific transfer RNA (tRNAPhe) from Saccharomyces. cerevisiae specifically between nucleotides D17 and G18 in the D-loop, with additional minor cleavages after D16 and G15. This makes lead(II) a sensitive structural probe for correct folding of tRNAPhe. In the present paper we use Pb2+ ions as a functional probe to determine whether this part of tRNA is protected by the Escherichia coli elongation factor EF-Tu in the ternary complex formed between Phe-tRNAPhe and EF-Tu.GTP. Our results show that for tRNA in complex with EF-Tu:GTP, the phosphodiester bond after D17 is cleaved, yet the phosphodiester bonds after D16 and G15 are not. To our knowledge, this is the first time that Pb2+ ions, bound at a specific site in tRNA, have been used both to investigate the correct folding of tRNA in complex, and to footprint a functional complex with components whose individual structures are known.
authors.affiliation: Department of Chemistry, Aarhus University, Denmark.
authors.forename: D E
authors.initials: DE
authors.lastname: Otzen
authors.forename: J
authors.initials: J
authors.lastname: Barciszewski
authors.forename: B F
authors.initials: BF
authors.lastname: Clark
mesh_terms: D001345:Autoradiography; D002845:Chromatography; D004926:Escherichia coli; D006160:Guanosine Triphosphate / Q000378:metabolism*; D007854:Lead / Q000378:metabolism*; D009690:Nucleic Acid Conformation; D010444:Peptide Elongation Factor Tu / Q000378:metabolism*; D012346:RNA, Transfer, Amino Acyl / Q000378:metabolism*; D012441:Saccharomyces cerevisiae / Q000378:metabolism
chemical_list: D012346:RNA, Transfer, Amino Acyl; C037801:tRNA, phenylalanylphenylalanine-; D007854:Lead; D006160:Guanosine Triphosphate; D010444:Peptide Elongation Factor Tu