Abstract: |
We describe the first direct observation of N-H···O=C hydrogen bonding in nucleic acids via the four-bond (4h)J(NjNj) coupling within an N1(i)-H1(i)···O6(j) = C6(j)-N1(j) segment of a G·G·G·G tetrad. The experiment, two-dimensional (2D) HN(N)-TOCSY, makes use of band-selective 15N isotropic mixing to transfer magnetization exclusively via the (4h)J(NN) couplings to yield correlations between N1H(i)(ω2) and N1(j)(ω1) across the hydrogen-bonded segment. In a complementary experiment, 2D cross-polarization (CP)-H(N)CO-(NN)TOCSY, employing band- selective heteronuclear 15N-13C cross-polarization sequences on either side of the 15N-15N TOCSY period, correlations are obtained between N1H(i)(ω2) and C6(j)(ω1) nuclei across the hydrogen bond. The symmetric A2X2-type coupling topology of the four-spin nitrogen system in the G·G·G·G tetrad permits accurate measurement of these couplings by a new procedure that fits the experimental data with known analytical isotropic mixing transfer functions. The techniques are demonstrated on a G·G·G·G tetrad formed within a novel dimeric quadruplex fold of a uniformly 13C/15N-labeled d(G-G-G-T-T-C-A-G-G) DNA sequence. The value of the (4h)J(NN) coupling constant for this system is estimated to be 0.136 ± 0.021 Hz. |