SOLUTION STRUCTURE OF AN OLIGODEOXYNUCLEOTIDE CONTAINING THE HUMAN N-RAS CODON-12 SEQUENCE REFINED FROM H-1-NMR USING MOLECULAR-DYNAMICS RESTRAINED BY NUCLEAR OVERHAUSER EFFECTS

The structure of d(GGCAGGTGGTG). d(CACCACCTGCC), consisting of codons 11, 12 (underlined), and 13 of the human n-ras protooncogene, was refi ned from H-1 NMR data. Patterns of internucleotide NOEs consistent wit h a B-form helix were observed for each strand. NOE intensities betwee n purine H8 and H1' protons were small compared to intensities between cytosine H5 and H6 protons, indicative of glycosyl torsion angles in the anti range. Cross-peaks were observed between purine H8 and pyrimi dine H5 and CH3 protons on adjacent bases in the direction of purine(5 '-->3')pyrimidine, but not in the direction pyrimidine( 5'-->3')purine . Watson-Crick hydrogen bonding between bases was intact. A total of 2 32 experimental distance restraints were obtained. Of these, 143 were intra-residue restraints and 89 were inter-residue restraints. A restr ained molecular dynamics/simulated annealing approach yielded 6 MD str uctures calculated from a B-form starting structure and 6 MD structure s from an A-form starting structure. These refined to an average pairw ise rms difference of 0.92 Angstrom, with maximum pairwise rmsd of 1.3 5 Angstrom. Accuracy of the emergent structures was assessed by relaxa tion matrix back-calculation. The sixth-root residual index of 7.0 x 1 0(-2) measured between the refined structures and the NOE intensity da ta suggested that the former were in reasonable agreement with the NOE data. The refined solution structures were in the B-family. Similar t o the human n-ras codon 61 sequence [Feng, B., & Stone, M. P. (1995) C hem. Res. Toxicol. 8, 821-832], the ras12 sequence contained local var iations in B-like conformation which did not confer large structural a lterations upon the duplex, but perhaps modulated the reactivity of th e first as compared to the second guanine in codon 12.