Sequential resonance assignment of the 500‐MHz nmr spectrum of d‐(CG)₆ and its structure under low‐salt conditions

Abstract

SynopsisTwo-dimensional (2D) nmr methods (correlated spectroscopy, nuclear Overhauser enhancement spectroscopy, and relayed correlated spectroscopy) have been used to obtain resonance assignment of the nonexchangeable base and sugar protons of a double-helical DNA segment, d-(CG), in D,O solutions under conditions of low ionic strength. Detailed information about the glycosidic torsion angle, sugar geometry, stacking patterns of the bases, and the overall solution structure of the dodecanucleotide has been obtained from the relative intensities of cross-peaks in the 2D spectra. The molecule shows general features of B-DNA under the experimental conditions employed. However, in spite of the repeating base sequence, there are subtle and detectable variations in the structure along the double helix. The terminal residues show considerable conformational flexibility.

Resonance assignments of non-exchangeable protons in B type DNA oligomers, an overview

“…It is noted in passing that this is typical of H2 protons: they apparently are so susceptible to long-range effects that the straightforward approach as in Fig 2 and Fig 3 does not work at all for these protons. Similar deviations in chemical shifts of protons belonging to the central unit of identical tri-nucleotides, measured under identical conditions, were also found for CGCGCGCGCGCG (18), AAAAAATTTTTT (6) and ATATATATAT (9). In fact, resonance assignments would have been virtually impossible for such sequences were it not for these small long-range effects.…”

Solution conformation of d-CTCGAGCTCGAG by two-dimensional NMR: conformational heterogeneity at XhoI cleavage site

“…The sugar geometries show wide variations ranging from C2,-endo to 04'-endo in contrast to the C2'-endo geometry implicated in regular B structure. This is in contrast to a regular structure observed in d-CGCGCGCGCGCG (Sheth et al, 1987b) under low-salt conditions. In addition, important structural variations at the cleavage sites of restriction enzymes BamHl (molecule I) and…”

Sequence-specific solution structure of d-GGTACGCGTACC

“…The other five J values are three-bond vicinal couplings which show very strong dependence on the conformation of the deoxyribose ring. The conformational dependence of these J values has been established [see, for example, Chary et al, (1987), Hosur et al (1986a,b), Sheth et al (1987b, and Rinkel and Aliona (1987)]. Table II lists expected J values for some selected conformations for the convenience of the discussion in this paper, along with the experimental results.…”

“…The NMR data from our own laboratories for the molecules I-V listed in the introduction and those reported by others also indicate that the B-DNA can adopt much wider sequence-dependent conformational variations. In solutions, average sugar geometry is closer to 04'-endo than to C2'-endo (Hosur et al, 1986a(Hosur et al, ,b, 1987bSheth et al, 1987a,b). The variations in the structure are not localized to the sugar pucker but are reflected in the other torsional angles, thereby altering the orientation of the phosphate groups and the relative positioning and stacking of the base pairs, which are the two important functional groups from the view of protein recognition.…”

“…The methodologies are being improved to obtain data of better quality (Hosur et al, 1985a(Hosur et al, ,b, 1987a, and new strategies are being developed to obtain more precise and detailed structural information from the experimental spectra. Low-resolution structures have been obtained earlier for the following double-helical DNA segments: (I) d-GGATCCGGATCC (Hosur et al, 1985;Ravikumar et al, 1985); (II) d-GAATTCGAATTC (Chary et al, 1987;Hosur et al, 1986a); (III) d-CTCGAGCTCGAG (Sheth et al, 1987a); (IV) d-GAATTCCCGAATTC (Hosur et al, 1986b); (V) d-(CG)6 (Sheth et al, 1987b). Among these, I-IV have recognition sites for the restriction enzymes BamUl, EcoRl, Xhol, and EcoRl, respectively.…”