NMR and UV studies of 3'-S-phosphorothiolate modified DNA in a DNA : RNA hybrid dodecamer duplex; implications for antisense drug design.
High-resolution NMR spectroscopy has been used to establish the conformational consequences of the introduction of a single 3[prime or minute]-S-phosphorothiolate link in the DNA strand of a DNA : RNA hybrid. These systems are of interest as potential antisense therapeutic agents. Previous studies on similarly modified dinucleotides have shown that the conformation of the sugar to which the sulfur is attached shifts to the north (C(3[prime or minute])-endo/C(2[prime or minute])-exo). Comparisons made between NOESY cross-peak intensities, and coupling constants from PE-COSY spectra, for both non-modified and modified duplexes confirm that this conformational shift is also present in the double helical oligonucleotide system. In addition it is noted that in both the dinucleotides and the modified duplex, the conformation of the sugar ring 3[prime or minute] to the site of modification is also shifted to the north. That this pattern is observed in the small monomeric system as well as the larger double helix is suggestive of some pre-ordering of the sequences. The conclusion is supported by consideration of the (1)H chemical shifts of the heterocyclic bases near the site of the modification. The enhanced stability that these conformational changes should bring was confirmed by UV thermal melting studies. Subsequently a series of singly and doubly 3[prime or minute]-S-phosphorothiolate-modified duplexes were investigated by UV. The results are indicative of an additive effect of the modification with thermodynamic benefit being derived from alternate spacing of two modified linkers.
Beevers
Andrew P G
AP
Department of Chemistry, University of Leeds, Leeds, UKLS2 9JT.
Fettes
Kevin J
KJ
Sabbagh
Ghalia
G
Murad
Fatima K
FK
Arnold
John R P
JR
Cosstick
Richard
R
Fisher
Julie
J
D001483:Base Sequence; D004247:DNA / Q000737:chemistry*; D015195:Drug Design; D009682:Magnetic Resonance Spectroscopy; D008969:Molecular Sequence Data; D009692:Nucleic Acid Heteroduplexes / Q000737:chemistry; D016376:Oligonucleotides, Antisense / Q000737:chemistry*; D010710:Phosphates / Q000737:chemistry*; D012313:RNA / Q000737:chemistry*; D013056:Spectrophotometry, Ultraviolet; D013816:Thermodynamics
C465503:3'-S-phosphorothiolate; D009692:Nucleic Acid Heteroduplexes; D016376:Oligonucleotides, Antisense; D010710:Phosphates; D012313:RNA; D004247:DNA