2003 Zweckstetter and Bax 2001).įor proteins, NMR structure determination is predominantly based on inter-proton NOEs. The alignment typically requires supplementation of NMR buffers with some type of alignment media, such as bicelles, nonionic polymers, Pf1 bacteriophages, anisotropically compressed gels or covalent modifications of investigated molecules with paramagnetic tags (Bax and Tjandra 1997 Clore et al. The direct spin–spin interactions can be measured under conditions where the studied molecules are at least partially aligned with respect to the magnetic field. In addition to these two sources, direct spin–spin interactions (D), known as (residual) dipolar couplings (RDCs), reveal the relative orientations of inter-nuclear vectors with respect to the direction of the external magnetic field. The major sources of structural information from NMR measurements of biomolecules in isotropic solution are nuclear Overhauser enhancements (NOEs), which provide information about short (<5 Å) inter-proton distances, and indirect spin–spin interactions that are characterized by scalar coupling constants (J), which provide information about torsion angles (Roberts 1993 Wijmenga and van Buuren 1998). We show that if the non-zero D contributions to J are not properly accounted for, they might cause structural artifacts/bias in NA studies that use solution NMR spectroscopy. For these couplings, the magnetic field-induced dipolar contributions were found to exceed the typical experimental error in J-coupling determinations by a factor of two or more and to produce considerable over- or under-estimations of the J coupling-related torsion angles, especially at magnetic field strengths >12 T and for NA fragments longer than 12 bp. We identified two classes of Js, namely 1J CH and 3J HH couplings, whose quantitative interpretation is notably biased by NA motional anisotropy. Here, we calculated the field-induced D contributions to 33 structurally relevant scalar coupling constants as a function of magnetic field strength, temperature and NA fragment size. This motional anisotropy is responsible for non-zero D contributions to Js. However, in strong magnetic fields, such as those employed for NMR analysis, the tumbling of NA fragments is anisotropic because the inherent magnetic susceptibility of NAs causes an interaction with the external magnetic field. In biomolecular NMR studies, it is commonly presumed that the dipolar contributions to Js are effectively canceled due to random molecular tumbling. The Hamiltonians for the D and J interactions have the same functional form thus, the experimentally measured apparent spin–spin coupling constant corresponds to a sum of J and D. Heteronuclear and homonuclear direct (D) and indirect (J) spin–spin interactions are important sources of structural information about nucleic acids (NAs).
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