Observation and measurement of internucleotide 2JNN coupling constants between 15N nuclei with widely separated chemical shifts.
Scalar coupling correlations between hydrogen bonded 15N nuclei in non Watson-Crick base pairs is a critical step in the structure determination of unusual nucleic acids. For observing the 2JNN coupling constant between far upfield N2,N6 (amino) nitrogens and far downfield (N1,N3,N7) nitrogens (separated by 150-160 ppm), the HNN-COSY experiment (Dingley and Grzesiek, 1998) is rather insensitive, due to technical difficulties associated with simultaneous excitation of both extremes of the 15N spectrum. These nuclei may be correlated by treating them in a pseudo-heteronuclear manner, using 15N selective pulses. The wide chemical shift separation allows accurate measurement of the 2JNN coupling constant using spin-echo difference methods. Pulse sequences for observation and measurement of 2JNN coupling constants between amino and N7 nuclei are presented and demonstrated on an A-A mismatch segment of the uniformly (15N,13C) labelled DNA sample, d(GGAGGAT)2.
Majumdar
A
A
Cellular Biochemistry and Biophysics Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA. majumdar@sbnmr1.ski.mskcc.org
Kettani
A
A
Skripkin
E
E
D020029:Base Pairing; D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D006860:Hydrogen Bonding; D019906:Nuclear Magnetic Resonance, Biomolecular / Q000379:methods; D009690:Nucleic Acid Conformation; D009838:Oligodeoxyribonucleotides / Q000737:chemistry*
D009838:Oligodeoxyribonucleotides; D004247:DNA