EFFECT OF DISTORTIONS IN THE PHOSPHATE BACKBONE CONFORMATION OF 6 RELATED OCTANUCLEOTIDE DUPLEXES ON CD AND P-31 NMR-SPECTRA

We examined the structural properties of six octanucleotide duplexes, d(TGACGTCA), d(ACTGCAGT), d(CTTCGAAG), d(CATCGATG), d(GTACGTAC), and d (CATGCATG). Circular dichroism (CD) and 2D P-31 and H-1 NMR spectrosco pies were used in conjunction. Although of the B-DNA type, it was poss ible to arrange CD spectra into two families, A and B. Family A resemb led poly(dG-dC) with a positive signal at approximately 280 nm and a n egative one at approximately 260 nm, while family B resembled poly(dA- dT) with a positive signal at approximately 270 nm and a negative one at approximately 250 nm. All P-31 resonances were assigned through con stant-time heteronuclear P-31-H-1 correlated spectra. J(H3'-P) couplin g constants related to dihedral angles epsilon (C4'-C3'-O3'-P) were de termined from H-1-P-31 J-resolved selective proton-flip 2D experiments . A good correlation was observed between P-31 chemical shifts and cou pling constants for all oligonucleotides. The patterns of these two pa rameters vs the base position along the sequences were almost similar. They were confronted with CD spectra. The results indicated that the position and magnitude of the signals were mainly affected by the CpG and ApT steps whose P-31 chemical shifts were the farthest away from t he mean P-31 chemical shift value. This is in keeping with greater rig idity at these steps and should explain the influence of the local ord er on the shape of the CD spectra. Lastly, both UV absorption and P-31 chemical shifts vs temperature provided normal temperature melting (T (m)) values for all of the octanucleotide duplexes except for d(CTTCGA AG), for which the T(m) was approximately 10-degrees-C lower compared to its counterpart d(CATCGATG). The decrease in the thermal stability of this octanucleotide duplex was imputed to its contained TT and AA r epeats, which might be able to induce correlated base destacking and p hosphate group distortion in the oligonucleotide and especially on the intermediate CpG. We demonstrate that the CpG step displayed P-31 NMR properties similar to those found in mismatched nucleotides exclusive ly in the d(CTTCGAAG) duplex.