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E DeJong

Publications and source records attributed to E DeJong.

3 recordsLinked to original sources
PubMed1997

Comparison of H5 and H8 relaxation rates of a 2H/13C/15N labeled RNA oligonucleotide with selective protonation at C5 and C8.

Uniformly 13C/15N enriched ribonucleotide monophosphates have been prepared with extensive deuterium enrichment of the non-exchangeable positions. The purine C8 and pyrimidine C5 base positions were selectively protonated prior to incorporation of the individual nucleotide triphosphates into an RNA oligonucleotide. The longitudinal and transverse relaxation rates of the H8 and H5 resonances of this deuterated molecule were compared with the relaxation rates of the corresponding protonated, 13C/15N enriched RNA molecule. Deuteration disrupts the efficiency of 1H-1H dipolar relaxation and reduces the longitudinal and transverse magnetization relaxation rates on average to 25% and 68%, respectively, of the values measured for the non-deuterated RNA molecule. Importantly, the longitudinal relaxation rates remain sufficiently rapid (> 1s[-1]) to permit the use of short recovery delays in multidimensional NMR experiments without significant loss of sensitivity.

Carbon Isotopes↗
PubMed1997

Preparation and characterization of a uniformly 2 H/ 15 N-labeled RNA oligonucleotide for NMR studies.

An RNA oligonucleotide that contains the binding site for Escherichia coli ribosomal protein S8 was prepared with uniform 15N isotopic enrichment and uniform deuterium enrichment at all non-exchangeable sites using enzymatic methods. The RNA binding site, which contains 44 nt, forms a hairpin in solution and requires Mg2+for proper folding. The longitudinal magnetization recovery rates of the exchangeable protons were compared for the [2H,15N]-enriched RNA molecule and for the corresponding fully [2H,15N]-enriched RNA hairpin. It was found that 1H-1H dipolar relaxation significantly contributes to the recovery of exchangeable proton longitudinal magnetization. The exchangeable proton resonance line widths were less affected by deuteration, indicating that chemical exchange with H2O remains the dominant mechanism of transverse magnetization relaxation. Nevertheless, deuteration of this RNA hairpin was found to enhance the sensitivity of NOE-based experiments relative to the fully protonated hairpin and to simplify 2D NMR spectra. The increased signal-to-noise ratio facilitated the assignment of the cytidine amino resonances and several of the purine nucleotide amino resonances and permitted the identification of NOE crosspeaks that could not be observed in spectra of the fully protonated RNA hairpin.

Binding Sites↗