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- Glemarec, C, et al.
(author)
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The NMR structure of 31-mer RNA domain of E. coli RNase P RNA using its non-uniformly deuterium labelled counterpart (the "NMR-window" concept)
- 1996
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In: Nucleic Acids Research. - : Oxford University Press (OUP). - 0305-1048 .- 1362-4962. ; 24:11, s. 2022-2035
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Journal article (peer-reviewed)abstract
- The NMR structure of a 31mer RNA constituting a functionally important domain of the catalytic RNase P RNA from Escherichia coli is reported. Severe spectral overlaps of the proton resonances in the natural 31mer RNA (1) were successfully tackled by unique spectral simplifications found in the partially-deuterated 31 mer RNA analogue (2) incorporating deuterated cytidines [C5 (>95 atom % 2H), C2' (>97 atom % 2H), C3' (>97 atom % 2H), C4' (>65 atom % 2H) and C5' (>97 atom % 2H)] [for the 'NMR-window' concept see: Földesi,A. et al. (1992) Tetrahedron, 48, 9033; Foldesi,A. et al. (1993) J. Biochem. Biophys. Methods, 26, 1; Yamakage,S.-I. et al. (1993) Nucleic Acids Res., 21, 5005; Agback,P. et al. (1994) Nucleic Acids Res., 22, 1404; Földesi,A. et al. (1995) Tetrahedron, 51, 10065; Földesi,A. et al. (1996) Nucleic Acids Res., 24, 1187-1194]. 175 resonances have been assigned out of total of 235 non-exchangeable proton resonances in (1) in an unprecedented manner in the absence of 13C and 15N labelling. 41 out of 175 assigned resonances could be accomplished with the help of the deuterated analogue (2). The two stems in 31mer RNA adopt an A-type RNA conformation and the base-stacking continues from stem I into the beginning of the loop I. Long distance cross-strand NOEs showed a structured conformation at the junction between stem I and loop I. The loop I-stem II junction is less ordered and shows structural perturbation at and around the G11 -C22 base pair.
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| 2. |
- Herrmann, Björn, et al.
(author)
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Differentiation of Chlamydia spp by sequence determination and restriction endonuclease cleavage of RNase P RNA genes
- 1996
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In: Journal of Clinical Microbiology. - 0095-1137 .- 1098-660X. ; 34:8, s. 1897-1902
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Journal article (peer-reviewed)abstract
- The amplification of DNA from Chlamydia trachomatis by PCR with degenerated primers yielded a 345-bp fragment of the putative RNase P RNA gene. From the deduced DNA sequence of this gene in C. trachomatis, a modified primer pair was designed. The primer pair was subsequently used to obtain the corresponding gene products from Chlamydia pneumoniae and Chlamydia psittaci. Sequence comparisons revealed similarities of 76.6% between C. trachomatis and C. pneumoniae, 79.5% between C. trachomatis and C. psittaci, and 84.7% between C. pneumoniae and C. psittaci. Furthermore, the three species were differentiated by fragment length polymorphism analysis after restriction enzyme cleavage of the PCR products. Sequence variations among 14 serotypes of C. trachomatis were confined to one purine base substitution in the putative RNase P RNA gene of lymphogranuloma venereum strains L1 to L3. Complete sequence similarity was found for nine strains of C. pneumoniae of different geographic origins. Taken together, our results indicate a possibility of the general application of this method in clinical bacteriology. Analysis of the secondary structures of the putative RNase P RNA genes from the different Chlamydia species suggested that a novel structural element in the domain of RNase P RNA is involved in base pairing with the 3'-terminal CCA motif of a tRNA precursor. This structure has not previously been found among RNase P RNAs of members of the division Bacteria.
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| 5. |
- Mikkelsen, Nils E, et al.
(author)
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Inhibition of RNase P cleavage by aminoglycosides
- 1999
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In: Proceedings of the National Academy of Sciences of the United States of America. - 0027-8424 .- 1091-6490. ; 96:11, s. 6155-6160
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Journal article (peer-reviewed)abstract
- A number of aminoglycosides have been reported to interact and interfere with the function of various RNA molecules. Among these are 16S rRNA, the group I intron, and the hammerhead ribozymes. In this report we show that cleavage by RNase P RNA in the absence as well as in the presence of the RNase P protein is inhibited by several aminoglycosides. Among the ones we tested, neomycin B was found to be the strongest inhibitor with a Ki value in the micromolar range (35 microM). Studies of lead(II)-induced cleavage of RNase P RNA suggested that binding of neomycin B interfered with the binding of divalent metal ions to the RNA. Taken together, our findings suggest that aminoglycosides compete with Mg2+ ions for functionally important divalent metal ion binding sites. Thus, RNase P, which is an essential enzyme, is indeed a potential drug target that can be used to develop new drugs by using various aminoglycosides as lead compounds.
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| 6. |
- Tallsjö, Annika, et al.
(author)
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Interaction between Escherichia coli RNase P RNA and the discriminator base results in slow product release
- 1996
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In: RNA. - 1355-8382 .- 1469-9001. ; 2:4, s. 299-307
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Journal article (peer-reviewed)abstract
- We suggested previously that a purine at the discriminator base position in a tRNA precursor interacts with the well-conserved U294 in M1 RNA, the catalytic subunit of Escherichia coli RNase P. Here we investigated this interaction and its influence on the kinetics of cleavage as well as on cleavage site selection. The discriminator base in precursors to tRNA(Tyr)Su3 and tRNA(Phe) was changed from A to C and cleavage kinetics were studied by wild-type M1 RNA and a mutant M1 RNA carrying the compensatory substitution of a U to a G at position 294 in M1 RNA. Our data suggest that the discriminator base interacts with the residue at position 294 in M1 RNA. Although product release is a rate-limiting step both in the absence and in the presence of this interaction, its presence results in a significant reduction in the rate of product release. In addition, we studied cleavage site selection on various tRNA(His) precursor derivatives. These precursors carry a C at the discriminator base position. The results showed that the mutant M1 RNA harboring a G at position 294 miscleaved a wild-type tRNA(His) precursor and a tRNA(His) precursor carrying an inosine at the cleavage site. The combined data suggest a functional interaction between the discriminator base and the well-conserved U294 in M1 RNA. This interaction is suggested to play an important role in determining the rate of product release during multiple turnover cleavage of tRNA precursors by M1 RNA as well as in cleavage site selection.
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