title: Resolution-optimized NMR measurement of (1)D(CH), (1)D(CC) and (2)D(CH) residual dipolar couplings in nucleic acid bases.
abstract: New methods are described for accurate measurement of multiple residual dipolar couplings in nucleic acid bases. The methods use TROSY-type pulse sequences for optimizing resolution and sensitivity, and rely on the E.COSY principle to measure the relatively small two-bond (2)D(CH) couplings at high precision. Measurements are demonstrated for a 24-nt stem-loop RNA sequence, uniformly enriched in (13)C, and aligned in Pf1. The recently described pseudo-3D method is used to provide homonuclear (1)H-(1)H decoupling, which minimizes cross-correlation effects and optimizes resolution. Up to seven (1)H-(13)C and (13)C-(13)C couplings are measured for pyrimidines (U and C), including (1)D(C5H5), (1)D(C6H6), (2)D(C5H6), (2)D(C6H5), (1)D(C5C4), (1)D(C5C6), and (2)D(C4H5). For adenine, four base couplings ((1)D(C2H2), (1)D(C8H8), (1)D(C4C5), and (1)D(C5C6)) are readily measured whereas for guanine only three couplings are accessible at high relative accuracy ((1)D(C8H8), (1)D(C4C5), and (1)D(C5C6)). Only three dipolar couplings are linearly independent in planar structures such as nucleic acid bases, permitting cross validation of the data and evaluation of their accuracies. For the vast majority of dipolar couplings, the error is found to be less than +/-3% of their possible range, indicating that the measurement accuracy is not limiting when using these couplings as restraints in structure calculations. Reported isotropic values of the one- and two-bond J couplings cluster very tightly for each type of nucleotide.
authors.affiliation: National Institute of Diabetes and Digestive and Kidney Diseases, National Institutes of Health, Laboratory of Chemical Physics, Bethesda, MD 20892-0520, USA.
authors.forename: Jérôme
authors.initials: J
authors.lastname: Boisbouvier
authors.forename: David L
authors.initials: DL
authors.lastname: Bryce
authors.forename: Erin
authors.initials: E
authors.lastname: O'neil-Cabello
authors.forename: Edward P
authors.initials: EP
authors.lastname: Nikonowicz
authors.forename: Ad
authors.initials: A
authors.lastname: Bax
mesh_terms: D020029:Base Pairing; D002247:Carbon Isotopes / Q000737:chemistry; D006147:Guanine / Q000737:chemistry*; D009587:Nitrogen Isotopes / Q000737:chemistry; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D010759:Phosphorus Isotopes / Q000737:chemistry; D011743:Pyrimidines / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*
chemical_list: D002247:Carbon Isotopes; D009587:Nitrogen Isotopes; D010759:Phosphorus Isotopes; D011743:Pyrimidines; D006147:Guanine; D012313:RNA