A Sensitive Multispectroscopic Probe for Nucleic Acids

Abstract

Azides have recently been used as vibrational probes of proteins, but their incorporation into nucleic acids has been limited to photo-cross-linking or click chemistry applications. The utility of 2'-azido-2'-deoxyuridine (N(3)-dU, 1) as an IR and (15)N NMR spectroscopic probe of the sugar phosphate backbone region of nucleic acids was investigated by measuring the effects of solvent, heterodimer formation, and temperature on peak frequencies and IR bandwidth. The azide IR asymmetric stretching band (nu(N(3))) of N(3)-dU was sensitive to its environment, undergoing a blue shift of 13.5 cm(-1) when changing the solvent from THF to water. The solvent effects on (15)N chemical shifts (delta((15)N)) of each of the nitrogen atoms in the azido group was studied, and the terminal nitrogen atom was the most sensitive to solvent, shifting downfield by 3.8 ppm when changing the solvent from THF-d(8) to D(2)O. Formation of a base-pair-like heterodimer between 3 (a silyl ether analogue of 1) and 2,6-diheptanamidopyridine (4) in chloroform resulted in minimal changes in the IR and (15)N NMR spectral frequency and chemical shift, respectively, as expected given the location of the azido moiety. The intrinsic temperature dependence of nu(N(3)) and delta((15)N) were found to be minimal over the temperature range studied especially compared to the solvent dependence of these spectral observables. The analysis of the experimental studies was complemented by density functional theory (DFT) calculations on model systems.

Two-Dimensional Infrared Study of Vibrational Coupling between Azide and Nitrile Reporters in a RNA Nucleoside

“…7, 12 The presence of these two reporters within a close proximity in the same molecule results in significant vibrational coupling between their dipoles. As a result, the relative intensities of the infrared transitions are sufficiently different than expected from prior literature 7, 12, 28 as well as the calculated transition dipole ratio. The observed vibrational frequencies and relative intensities of the linear IR spectrum can often be predicted from the solutions to the Hamiltonian shown below: $$H=true(\begin{array}{cc}left{\epsilon }_{N_3}& left{\beta }_{12}\\ left{\beta }_{12}& left{\epsilon }_{CN}\end{array}true),$$ where β 12 represents the coupling between the azide (ε N 3 ) and nitrile (ε CN ) vibrational modes.…”

“…21, 24 Further studies have extended the application of these probes in nucleosides to infrared, 2D IR, and NMR experiments. 28, 29 The sensitivity of these reporters to the local environment 1, 2, 8, 28 and hydrogen bonding 2, 5 provides an avenue to track these dynamics by 2D IR methods using chemical exchange 30, 31 and spectral diffusion. 11–13, 17 …”

“…3, 28, 29, 42, 46 With these synthetic approaches, N 3 and CN probes can be placed virtually anywhere providing a useful monitor of site-specific dynamics in environments such as the major groove, minor groove, and phosphate-sugar region. Herein, we demonstrate the effectiveness of a pair of vibrational reporters as tools to investigate structural and dynamic aspects of different regions of DNA and RNA aptamers using 2D IR.…”

Two-dimensional infrared study of 3-azidopyridine as a potential spectroscopic reporter of protonation state

“…Recent work from the groups of both Cho and Brewer [42][43][44] indicate the potential for azido-derivatized compounds as vibrational probes. Organic azides are particularly interesting because they tend to have much larger transition moments than nitriles and thiocyanates.…”

Applications of nuclear shielding

“…297 The utility of 2 0 -azido-2 0 -deoxyuridine (N3-dU, 1) as an IR and 15 N NMR spectroscopic probe of the sugar phosphate backbone region of nucleic acids is investigated by measuring the effects of solvent, heterodimer formation and temperature on peak frequencies and IR bandwidth. 298 Various [5,6]pyrano[2,3-c]pyrazol-4( 1 H)-thiones were synthesised in high yields by treatment of the corresponding [5,6]pyrano[2,3-c]pyrazol-4( 1 H)-ones with Lawesson's reagent. Complete and unambiguous assignment of chemical shifts ( 1 H, 13 C, 15 N) and coupling constants ( 1 H, 1 H; 13 C, 1 H) was achieved by the combined application of various one-and two-dimensional (1D and 2D) NMR spectroscopic techniques.…”