5′-CGA sequence is a strong motif for homo base-paired parallel-stranded DNA duplex as revealed by NMR analysis

The structure of the non-self-complementary DNA heptamer d(CGACGAC) at low pH has been determined by the quantitative NMR refinement procedure designated SPEDREF (SPEctral-Driven REFinement). Acid-base titration of the molecule indicated a prominent n = 2 pK_a near 6.8. In the pH range up to 6.0, the heptamer forms a remarkably stable double helix, which was conclusively shown to be an unusual homobase-paired parallel-stranded double helix (termed II-DNA). In this II-DNA helix, the 5′-CGA trinucleotide is the structural motif that accounts for the stability, with the C⁺·C hemiprotonated base pair (in which C⁺ is N³-protonated cytosine) providing for the alignment site and the unusual interstrand G-A base stack in the GpA step furnishing the additional stabilizing forces. The exchangeable proton data from two-dimensional nuclear Overhauser effect spectroscopy are in total agreement with the refined structure. We conclude that the 5′-CGA or other related sequences (e.g., 5′-CCGA) are powerful motifs in promoting the II-DNA or II-RNA conformations that may play certain biological functions.