Hydrogen-1 and phosphorus-31 NMR studies of oligonucleotides directed towards investigating the sequence-dependent structural nature of DNA
Abstract
Assignments of the $\sp{31}$P resonances of a series of six sequence-related tetradecamer and two dodecamer DNA duplexes were determined by either site-specific $\sp{17}$O labelling of the phosphoryl groups or by two-dimensional $\sp1$H-$\sp{31}$P heteronuclear NMR methods. Comparisons of the $\sp{31}$P resonances of these sequences have allowed greater insight of various factors responsible for $\sp{31}$P chemical shifts in oligonucleotides. Common patterns of the $\sp{31}$P shift variation along the DNA duplex are shown to be sequence-dependent. The observed variation is largely a function of the purine-pyrimidine base step arrangement of the sequence. An accumulation of the $\sp{31}$P shift values from these sequences, as well as others presented in the literature allowed the formation of $\sp{31}$P shift rules, which can be used to predict the $\sp{31}$P NMR spectrum of other oligonucleotide sequences. Comparison of the $\sp{31}$P shift variation to variations in DNA helical geometry obtained from crystallographic studies of oligonucleotides in the literature indicates a sequence-dependent structural relationship exists. The $\sp{31}$P shifts correlate well with some geometric helical parameters, as predicted by the Dickerson/Calladine sum function rules. These correlations are consistent with the hypothesis that the structural variation is responsible for perturbing the deoxyribose phosphate backbone, which responds to changes in local helical structure. The variation of the $\sp{31}$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. Inter- and intra- nucleotide $\sp1$H-$\sp1$H distances were determined for four sequence-related tetradecamer sequences using a series of time-developed nuclear Overhauser enhancement measurements. Comparison between sets of distances verified changes in helical conformation occur due to single base pair changes in each sequence, which is consistent with the predicted sequence-dependent model of DNA. The combined results of the $\sp{31}$P chemical shift and interproton distance determinations demonstrate that sequence-specific structural variations exist along the DNA double helix in solution. These variations, however, are not necessarily identical to the structural changes observed in x-ray crystallographic studies of oligonucleotides.
Degree
Ph.D.
Advisors
Gorenstein, Purdue University.
Subject Area
Biochemistry|Analytical chemistry|Chemistry
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