Two-dimensional¹H and³¹P NMR spectra and restrained molecular dynamics structure of an oligodeoxyribonucleotide duplex refined via a hybrid relaxation matrix procedure

Assignment of the¹H and³¹P resonances of a decamer DNA duplex, d(CGCTTAAGCG)₂ was determined by two-dimensional COSY, NOESY and¹H-³¹P Aire Absorption phase Constant time (PAC) heteronuclear correlation spectroscopy. The solution structure of the decamer was calculated by an iterative hybrid relaxation matrix method combined with NOESY-distance restrained molecular dynamics. The distances from the 2D NOESY spectra were calculated from the relaxation rate matrix which were evaluated from a hybrid NOESY volume matrix comprising elements from the experiment and those calculated from an initial structure. The hybrid matrix-derived distances were then used in a restrained molecular dynamics procedure to obtain a new structure that better approximates the NOESY spectra. The resulting partially refined structure was then used to calculate an improved theoretical NOESY volume matrix which is once again merged with the experimental matrix until refinement is complete J_H3’-P coupling constants for each of the phosphates of the decamer were obtained from ‘H- P J-resolved selective proton flip 2D spectra. By using a modified Karplus relationship the C4’-C3’-03’-P torsional angles (ε) were obtained. Comparison of the³¹P chemical shifts and J_H3’-P coupling constants of this sequence has allowed a greater insight into the various factors responsible for³¹P chemical shift variations in oligonucleotides. It also provides an important probe of the sequence-dependent structural variation of the deoxyribose phosphate backbone of DNA in solution. These correlations are consistent with the hypothesis that changes in local helical structure perturb the deoxyribose phosphate backbone. The variation of the 3IP chemical shift, and the degree of this variation from one base step to the next is proposed as a potential probe of local helical conformation within the DNA double helix. The pattern of calculated ε and ζ torsional angles from the restrained molecular dynamics refinement agrees quite well with the measured J_H3’-P coupling constants. Thus, the local helical parameters determine the length of the phosphodiester backbone which in turn constrains the phosphate in various allowed conformations.