1H-1H correlations across N-H•••N hydrogen bonds in nucleic acids

In (2)HJ(NN)-COSY experiments, which correlate protons with donor/acceptor nitrogens across N-d. . . HNa bonds, the receptor nitrogen needs to be assigned in order to unambiguously identify the hydrogen bond. For many situations this is a non-trivial task which is further complicated by poor dispersion of (N-a,N-d) resonances. To address these problems, we present pulse sequences to obtain direct, internucleotide correlations between protons in uniformly C-13/N-15 labeled nucleic acids containing N-d. . . HNa hydrogen bonds. Specifically, the pulse sequence H2(N1N3)H3 correlates H2(A,omega (1)):H3(U,omega (2)) protons across Watson-Crick A-U and mismatched G .A base pairs, the sequences H5(N3N1)H1/H6(N3N1)H1 correlate H5(C,omega (1))/H6(C,omega (1)):H1(G,omega (2)) protons across Watson-Crick G-C base pairs, and the H-2(N2N7)H8 sequence correlates NH2(G,A,C;omega (1)):H8(G,A;omega (2)) protons across G .G, A .A, sheared G .A and other mismatch pairs. These H-1-H-1 connectivities circumvent the need for independent assignment of the donor/acceptor nitrogen and related degeneracy issues associated with poorly dispersed nitrogen resonances. The methodology is demonstrated on uniformly C-13/N-15 labeled samples of (a) an RNA regulatory element involving the HIV-1 TAR RNA fragment, (b) a multi-stranded DNA architecture involving a G . (C-A) triad-containing G-quadruplex and (c) a peptide-RNA complex involving an evolved peptide bound to the HIV-1 Rev response element (RRE) RNA fragment.