Abstract
The DNA undecamers GTACAAAGTAC (AAA 11-mer) and GTACGAGGTAC (GAG 11-mer) have been studied in solution by high-resolution NMR spectroscopy. Both duplexes form stable hairpins containing single deoxyadenosine loops and stems containing five base-pairs that are closed at the loop end by sheared A·A and G·C pairs, respectively. These molecules thus contain new AAA and GAG loop turn motifs. All protons, including the chiral H5'/H5'' protons of the loop residues, were assigned using NOESY, DQF-COSY and heteronuclear 1 H- 31 P COSY experiments. The backbone torsion angles were constrained using experimental data from NOE crosspeaks, three-bond 1 H- 1 H coupling constants and four-bond 1 H- 31 P coupling constants and four-bond 1 H- 31 P coupling constants. The AAA and GAG 11-mers form similar structures in solution. The detailed structure of the AAA 11-mer was determined by the combined use of NMR, distance geometry and energy minimization methods. This structure exhibits good stacking of the loop adenosine base on the closing 5A·7A sheared pair, with the 6A base stacking on the 5A base and the 6A deoxyribose stacking with the 7A base. All sugars in the AAA 11-mer hairpin adopt the typical DNA C2'-endo conformation and a sharp backbone turn occurs between residues 6A and 7A. This loop turn is brought about mainly by a change in the backbone phosphate torsion angles from ζ(g - ) α(g - ) to ζ(g + ) α(g + ) at the turn. The γ torsion angle of residue 7A in the closing sheared pair also changes from gauche + to trans. In Pu 1 NPu 2 loop turns of the GCA, AAA and GAG types, the chemical shift of the H4' proton of the loop deoxyribose depends on the nature of Pu 2 ; this reflects the stacking of the loop sugar on the Pu 2 base and the different ring current effects of A or G in this position.