NMR SOLUTION STRUCTURE OF A DNA DODECAMER CONTAINING SINGLE G-CENTER-DOT-T MISMATCHES

The three-dimensional solution structure of the self-complementary DNA dodecamer [GRAPHICS] which contains the thermodynamically destabilizi ng [GRAPHICS] motif was determined using two-dimensional NMR spectrosc opy and simulated annealing protocols. Relaxation matrix analysis meth ods were used to yield accurate NOE derived distance restraints. Scala r coupling constants for the sugar protons were determined by quantita tive simulations of DQF-COSY cross-peaks and used to determine sugar p ucker populations. Twenty refined structures starting from random geom etries converged to an average pairwise root mean square deviation of 0.49 Angstrom. Back calculated NOEs give R-C and R-X factors of 0.38 a nd 0.088, respectively. The final structure shows that each of the sin gle G T mismatches form a wobble pair with two hydrogen bonds where th e guanine projects into the minor groove and the thymine projects into the major groove. The incorporation of the destabilizing [GRAPHICS] m otif has little effect on the backbone torsion angles and helical para meters compared to standard B-form duplexes, which may explain why G T mismatches are among the most commonly observed in DNA, The structure shows that perturbations caused by a G.T mismatch extend only to its neighboring Watson-Crick base pair, thus providing a structural basis for the applicability of the nearest-neighbor model to the thermodynam ics of internal G T mismatches.