Structural and Thermodynamic Studies of d(C3T4C3) in Acid Solution: Evidence for Formation of the Hemiprotonated CH+·C Base Pair

Daniel S. Pilch, Richard H. Shafer

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In an effort to probe the structure and stability of the CH+·C base pair in DNA, we have studied the conformation of the deoxydecanucleotide d(C3T4C3) at pH 5.5, using 1H-NMR and UV absorption spectroscopy. Lowering the pH of a solution containing d(C3T4C3) is accompanied by the induction of six new imino proton NMR resonances in a region (15.4–15.8 ppm) downfield from where either T-H3 or G-H1 Watson-Crick (W-C)-paired imino protons normally resonate. These novel resonances correspond to CH+-H3 imino protons engaged in hydrogen bonding with the N3 atoms of unprotonated cytosine residues. Since six, and not three, imino proton resonances arise upon lowering of the pH, the base-paired structure formed by d(C3T4C3) appears to be a parallel-stranded, bimolecular duplex [d(CH+3T4CH+3)·d(C3T4C3)], containing six CH+·C base pairs and an inner loop of eight non-base-paired thymine residues. A nuclear Overhauser enhancement spectroscopy (NOESY) experiment demonstrates dipolar contacts between the CH+-H3 imino protons and both CH+-H4 and C-H4 amino protons, confirming the presence of CH+·C base pairs. Thermal denaturation profiles at differing DNA concentrations are consistent with the existence of a bimolecular duplex at low pH. Thermodynamic analysis of UV melting curves indicates that CH+·C base pairs are more stable than either Watson-Crick G·C or A·T base pairs.

Original languageEnglish (US)
Pages (from-to)2565-2571
Number of pages7
JournalJournal of the American Chemical Society
Issue number7
StatePublished - Apr 1 1993
Externally publishedYes

All Science Journal Classification (ASJC) codes

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry


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