Solution Structure of two Mismatches A · A and T · T in the K‐ras Gene Context by Nuclear Magnetic Resonance and Molecular Dynamics

Two mismatches, one homopurine (A · A) and the other homopyrimidine (T · T), have been incorporated at the central position N of: 5′d(GCCACNAGCTC) · d(GAGCTN GTGGC) in order to study nuclear magnetic resonance spectra and molecular dynamics. These duplexes constitute the sequence 29–39 of the K‐ras gene coding for Gly12, a hot spot for mutation. The NMR spectra show that the duplexes are not greatly distorted by the introduction of the mismatches and their global conformation is that of a canonical B‐form double helix. For both systems, no structural change is observed in the pH range 4.7–9. For the duplex containing the homopurine A · A mismatch, we propose a type of pairing involving one hydrogen bond between the amino group of one central adenine and the nitrogen N1 of the opposite adenine. For the duplex containing the mispaired T · T bases, NMR spectra recorded in H₂O at 282 K indicate that these central bases are engaged in wobble pairing, involving two imino‐carbonyl hydrogen bonds. For both systems two conformations with the same donor and acceptor pattern can coexist, one being obtained from the other by a 180° rotation about the pseudodyadic axis. Exchange between the two forms is observed by NMR at low temperature for the T · T mispair and also inferred from NMR measurements on the A · A system. The presence of this exchange and its pathway has been investigated by molecular dynamics calculations on both systems. Distance restrained and unrestrained molecular dynamics are in very good agreement with the NMR data. The average structure for either mispair shows only small conformational change from normal B DNA. For each, a systematic pathway is observed for exchange between the two conformations.