HOBS methods for enhancing resolution and sensitivity in small DNA oligonucleotide NMR studies

Abstract

(1) H NMR spectra from biopolymers give chemical shifts classified according to proton type and often suffer from signal degeneracy. Data from nucleic acids are particularly prone to this failing. Recent developments in proton broadband decoupling techniques with the promise of enhanced resolution at full sensitivity have allowed us to investigate the application of homonuclear band-selective (HOBS) decoupling to the study of small synthetic DNA molecules and to compare these with results from classical and pure shift techniques. Improved signal resolution at full sensitivity in both HOBS-1D (1) H and HOBS-2D [(1) H, (1) H] NOESY NMR data is reported for three example small DNA molecules. Comparisons of (1) H T1 and integrals of signals from HOBS-1D (1) H and HOBS-2D [(1) H, (1) H] NOESY NMR data with those of standard data collection methods are also reported. The results show that homonuclear HOBS-NOESY data are useful for data assignment purposes and have some merit for quantification purposes. In general, we show that resolution and sensitivity enhancement of (1) H NMR data for small DNA samples may be achieved without recourse to higher magnetic field strength at full sensitivity in a band-selected manner.

Broadband1H homodecoupled NMR experiments: recent developments, methods and applications

“…An important advantage of the HOBS technique is its easy and reliable implementation for a large number of homonuclear and heteronuclear multidimensional experiments, as reported for HOBS‐NOESY, HOBS‐ROESY, HOBS‐TOCSY, HOBS‐HSQC, HOBS‐HSQMBC, HOBS‐inversion‐recovery (IR) and HOBS‐CPMG‐PROJECT, with particular applications for the measurement of heteronuclear coupling constants, enantiodifferentiation studies, discrimination of diastereoisomers and the measurement of T 1 and T 2 NMR relaxation times . In addition, it can be also attractive for biomolecular NMR applications, as reported for the effective elimination of residual HH dipolar couplings (RDCs) contributions to line broadening when working with partially oriented proteins in anisotropic media …”

“…A requirement for a success implementation of any ZS module is to have in‐phase (IP) HH magnetization, because signals involving anti‐phase (AP) components, like those found in conventional COSY or HMBC, cancel under homodecoupling conditions. The resulting pure shift spectra have a wide range of potential uses, as demonstrated for the analysis of diastereomeric or complex mixtures, to carry out structural elucidation studies, to analyze diffusion and molecular dynamic processes or to measure heteronuclear coupling constants …”

“…Thus, T 1 and T 2 relaxation times can be accurately measured from the resulting singlet lines using conventional mono‐exponential curve‐fitting methods. Examples have been provided for the undecapeptide cyclosporine, the small six‐mer oligonucleotide d(GCCTGC) and the 16‐mer d(CGACGCGTACGCGTCG)2 DNA duplex . Figure A shows the basic scheme of the 1D HOBS‐IR experiment where the conventional acquisition scheme has been replaced by an HOBS detection block.…”

perfectBASH: Band‐selective homonuclear decoupling in peptides and peptidomimetics

“…Initially developed for broadband BIRD homonuclear decoupling by Lupulescu and Frydman, real‐time decoupling variants using the Zangger–Sterk element and soft pulses for band‐selective decoupling were also presented. The band‐selective variant of the real‐time homonuclear decoupling (HOBS) has been implemented into homonuclear and heteronuclear 2D experiments as well as experiments for the measurement of relaxation times . Again, major applications of these band‐selective homodecoupling techniques are the analysis of α‐peptides and proteins .…”

“…Again, major applications of these band‐selective homodecoupling techniques are the analysis of α‐peptides and proteins . Nevertheless, applications in the analysis of oligosaccharides, small DNA fragments, or the determination of the enantiomeric and diastereomeric excess are also reported using band‐selective decoupled 1D and 2D NMR experiments. It is common to these BASH decoupling techniques that they can only be used successfully in the stated applications, if the proton region, which should be decoupled, is well defined and free of mutually coupled protons.…”