TWo-Dimensional¹H and³¹P NMR Spectra and Restrained Molecular Dynamics Structure of an Oligodeoxyribonucleotide Duplex Refined via a Hybrid Relaxation Matrix Procedure

Abstract

Assignment of the 1 H and 31 P resonances of a decamer DNA duplex, d(CGCTTAAGCG)2 was determined by two-dimensional COSY, NOESY, and 1 H-31 P Pure 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. JH3′-P coupling constants for each of the phosphates of the decamer were obtained from 1 H-31 P J-resolved selective proton flip 2D spectra. By using a modified Karplus relationship the C4′-C3′-O3′-P torsional angles (ε) were obtained. Comparison of the 31 P chemical shifts and JH3′-P coupling constants of this sequence has allowed a greater insight into the various factors responsible for 31 P 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 31 P 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 ( 1 9 9 0 ) 2 from the restrained molecular dynamics refinement agrees quite well with the measured JH3′-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.

DNA structural forms

“…This set of structures can be summarised by several general characteristics. Firstly, in contrast to crystallographic results, canonical phosphodiester backbone conformations dominate in solution and there is little evidence for either B n conformations (e£ = g~t) or ay crankshaft transitions (g"g + "tt) (Powers et al 1990;Karslake et al 1992;Kim & Reid, 1992;Hartmann et al 1993). Nevertheless, the backbone dihedrals are clearly seen to be variable, linked to base sequence and showing coupling with both sugar puckering and certain helicoidal parameters (Schmitz et al 1991;Karslake et al 1992;Kim & Reid, 1992;Schmitz et al 1993;El Antri et al 1993).…”

Conformational variation of the central CG site in d(ATGACGTCAT)2 and d(GAAAACGTTTTC)2

“…The 31 P spectra are referenced to trimethylphosphate at 0.0 p.p.m. The long‐range heteronuclear correlation (COLOC) [27–30] is acquired in 300/502 scans for each of the 128 free‐induction decays, which contain 2048 data points over 600/530 Hz spectral widths. Zero‐filling in f 1 produces 2048 × 1024 data matrices.…”

Conformation of the circular dumbbell d〈pCGC-TT-GCG-TT〉: Structure determination and molecular dynamics

“…phodiester torsion angles c~ and ~ (Gorenstein, 1984;Powers et al, 1990). Phosphorus resonances of the dumbbell were assigned previously by means of 2D 31p-1H correlated spectra (Ippel et al, 1995a).…”