NMR Studies of 3'-S-phosphorothiolate Modified Oligonucleotides

The structure-function relationship of nucleic acids is highly researched due to their , potential as therapeutics and bio-analytical tools. The structural effects of a subtle 3'-S-phosphorothiolate (3'-8) modification within DNA systems has been investigated here with regards to a role in antisense therapy. NMR spectroscopy was the technique of choice for this study, and was firstly used to' investigate the effect of 3'-8 modifications within a deoxytliymidine trinucleotide. Proton-proton coupling constants were extracted from this data and enabled a full conformational analysis of the system. The incorporation of a single modification induced a change in the conformation of the attached and 3' sugar ring (3' effect) from a south (DNA-like) towards a north (RNA-like) pucker. However, in the doubly modified trinucleotide although the sugar attached to the modification changed to a north pucker there was no evidence of the 3' sugar being effected. It appears that multiple alternate modifications may have benefits over multiple -- ------ ------. -----_._--------------------------- - -----------------. sequential modifications. NMR spectroscopy was then used to determine the structure of a DNA duplex and the effects of the 3'-8 modifications within this duplex. Two alternate modifications were incorporated into a standard B-type DNA helix and the conformational changes examined. Upon incorporation of the modifications, the sugars attached to the 3'-8 modifications underwent a conformational change from south-type sugar puckers to north-type puckers, and the sugars 3' to the modifications changed to a 50:50 north:south conformation. This was consistent to the effects of single modification within trlnucleotides and highlights the localised effects ofthis modification. UV spectroscopy was used to 'examine the stability of the duplexes. The changes observed with the 3'-8 modifications resulted in a destabilised DNA duplex, compared to the unmodified duplex. From the presence of the 3' effect within a DNA duplex, it was determined that the 3' effect is due to favourable intrastrand base stacking.EThOS - Electronic Theses Online ServiceGBUnited Kingdo