G.V. Fazakerley

Ionized and wobble base-pairing for bromouracil-guanine in equilibrium under physiological conditions: A nuclear magnetic resonance study on an oligonucleotide containing a bromouracil-guanine base-pair as a function of pH☆

A one and two-dimensional nuclear magnetic resonance study of a non-selfcomplementary oligonucleotide containing a central 5-bromouracil-guanine pair is reported. For these two bases three types of hydrogen bonding schemes could exist; wobble, rare tautomer and ionized. The two-dimensional spectra of non-exchangeable protons together with one-dimensional spectra recorded in water show that at pH 7.0 the predominant species is a right-handed B-form DNA in which the brU.G pair has wobble geometry. On raising the pH we observe a transition monitored by proton chemical shift changes for the brU.G and adjacent base-pairs. The mid-point of the transition was observed at pH 8.6. Spectra recorded at pH 9.8 show that the helix remains intact with B form conformation. It is shown that this high pH form has an ionized brU.G base-pair now in Watson-Crick geometry. Thus under physiological conditions an equilibrium exists between wobble and ionized structures.

Structural and dynamic properties of a bromouracil-adenine base pair in DNA studied by proton NMR

We have synthesized and studied by proton NMR a duplex heptaoligonucleotide containing a 5-bromouracil (brU)-adenine base pair. This represents the first structural characterization of a B-form DNA containing brU. The brU.A base pair is Watson-Crick rather than Hoogsteen as seen for the monomers in the crystalline state. From analysis of the NOESY sepctra at very short mixing times evidence is presented that substitution of brU for T induces significant conformational changes from that of a normal B DNA. The helix twist between brU4.A11 and G3.C12 is ca. 15 degrees and for both brU4 and G3 the glycosyl torsion angles are significantly changed. The imino proton of the bru.A base pair shows a pH insensitive line with which shows that the pK of brU in this base pair is very much higher than that of the monomer.