DNA base order parameter determination without influence of chemical exchange

Nuclear magnetic resonance (NMR) spectroscopy is a versatile tool used to investigate the dynamic properties of biological macromolecules and their complexes. NMR relaxation data can provide order parameters S², which represent the mobility of bond vectors reorienting within a molecular frame. Determination of S² parameters typically involves the use of transverse NMR relaxation rates. However, the accuracy in S² determination can be diminished by elevation of the transverse relaxation rates through conformational or chemical exchange involving protonation/deprotonation or non-Watson-Crick base-pair states of nucleic acids. Here, we propose an approach for determination of S² parameters without the influence of exchange processes. This approach utilizes transverse and longitudinal ¹³C chemical shift anisotropy (CSA) – dipole–dipole (DD) cross-correlation rates instead of ¹³C transverse relaxation rates. Anisotropy in rotational diffusion is taken into consideration. An application of this approach to nucleotide base CH groups of a uniformly ¹³C/¹⁵N-labeled DNA duplex is demonstrated.