ABSTRACT
The properties of gapmer antisense oligonucleotide (ASO) flanked by deoxyribonucleic guanidine (DNG) were investigated for the potential application in antisense technology. DNG is a unique nucleotide analog which has a positively charged internucleotide guanidinium linkage instead of negatively charged phosphodiester backbone linkage. We prepared a gapmer ASO containing DNG units at both wings of the sequence and compared its properties with 2',4'-BNA/LNA gapmer ASOs with phosphorothioate (PS) backbone. Although DNG gapmer showed no stabilizing effect on the duplex formation with target RNA, the DNG modification was found to be tolerant to exonuclease digestion. Furthermore, DNG gapmer can induce RNase H-mediated cleavage of target RNA molecule, a requisite property for the antisense strategy. Therefore, the DNG gapmer developed in this study could be an interesting and useful candidate for the development of potent ASOs.
Subject(s)
Guanidines/chemistry , Oligonucleotides, Antisense/chemistry , Oligonucleotides, Antisense/genetics , Base Sequence , Chemistry Techniques, Synthetic , DNA/chemistry , DNA/genetics , Humans , Molecular Structure , Oligonucleotides, Antisense/chemical synthesis , RNA/chemistry , RNA/genetics , RNA Cleavage , Thermodynamics , Transition TemperatureABSTRACT
The i-motif structures are formed by oligonucleotides containing cytosine tracts under acidic conditions. The folding of the i-motif under physiological conditions is of great interest because of its biological role. In this study, we investigated the effect of the intra-strand cross-link on the stability of the i-motif structure. The 4-vinyl-substituted analog of thymidine (T-vinyl) was incorporated into the 5'-end of the human telomere complementary strand, which formed the intra-strand cross-link with the internal adenine. The intra-strand cross-linked i-motif displayed CD spectra similar to that of the natural i-motif at acidic pH, which was transformed into a random coil with the increasing pH. The pH midpoint for the transition from the i-motif to random coil increased from pH 6.1 for the natural one to pH 6.8 for the cross-linked one. The thermodynamic parameters were obtained by measuring the thermal melting behaviors by CD and UV, and it was determined that the intra-strand cross-linked i-motif is stabilized due to a favorable entropy effect. Thus, this study has clearly indicated the validity of the intra-strand cross-linking for stabilization of the i-motif structure.
