Abstract:
:Characterization of the structure and dynamics of nucleic acids by NMR benefits significantly from position specifically labeled nucleotides. Here an E. coli strain deficient in the transketolase gene (tktA) and grown on glucose that is labeled at different carbon sites is shown to facilitate cost-effective and large scale production of useful nucleotides. These nucleotides are site specifically labeled in C1' and C5' with minimal scrambling within the ribose ring. To demonstrate the utility of this labeling approach, the new site-specific labeled and the uniformly labeled nucleotides were used to synthesize a 36-nt RNA containing the catalytically essential domain 5 (D5) of the brown algae group II intron self-splicing ribozyme. The D5 RNA was used in binding and relaxation studies probed by NMR spectroscopy. Key nucleotides in the D5 RNA that are implicated in binding Mg(2+) ions are well resolved. As a result, spectra obtained using selectively labeled nucleotides have higher signal-to-noise ratio compared to those obtained using uniformly labeled nucleotides. Thus, compared to the uniformly (13)C/(15)N-labeled nucleotides, these specifically labeled nucleotides eliminate the extensive (13)C-(13)C coupling within the nitrogenous base and ribose ring, give rise to less crowded and more resolved NMR spectra, and accurate relaxation rates without the need for constant-time or band-selective decoupled NMR experiments. These position selective labeled nucleotides should, therefore, find wide use in NMR analysis of biologically interesting RNA molecules.
journal_name
J Biomol NMRjournal_title
Journal of biomolecular NMRauthors
Thakur CS,Luo Y,Chen B,Eldho NV,Dayie TKdoi
10.1007/s10858-011-9586-1subject
Has Abstractpub_date
2012-02-01 00:00:00pages
103-14issue
2eissn
0925-2738issn
1573-5001journal_volume
52pub_type
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journal_title:Journal of biomolecular NMR
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journal_title:Journal of biomolecular NMR
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章,评审
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journal_title:Journal of biomolecular NMR
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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更新日期:2016-10-01 00:00:00
abstract::Proton chemical shifts of a series of disordered linear peptides (H-Gly-Gly-X-Gly-Gly-OH, with X being one of the 20 naturally occurring amino acids) have been obtained using 1D and 2D 1H NMR at pH 5.0 as a function of temperature and solvent composition. The use of 2D methods has allowed some ambiguities in side-chai...
journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
doi:10.1007/BF00227466
更新日期:1995-01-01 00:00:00
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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更新日期:2000-06-01 00:00:00
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journal_title:Journal of biomolecular NMR
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更新日期:2003-05-01 00:00:00
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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journal_title:Journal of biomolecular NMR
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journal_title:Journal of biomolecular NMR
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更新日期:2005-04-01 00:00:00
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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更新日期:2014-11-01 00:00:00
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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更新日期:2014-01-01 00:00:00
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journal_title:Journal of biomolecular NMR
pub_type: 杂志文章
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journal_title:Journal of biomolecular NMR
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abstract::The arrival of very high field magnets and cryogenic circuitries, and the development of relaxation-optimized pulse sequences have added powerful tools for increasing sensitivity and resolution in NMR studies of biomacromolecules. The potential of these advances is not fully realized in practice, however, since curren...
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pub_type: 杂志文章,评审
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