Structure and Stability of the Consecutive Stereoregulated Chiral Phosphorothioate DNA Duplex

Abstract

The duplex structures of the stereoregulated phosphorothioate DNAs, [R(p),R(p)]- and [S(p),S(p)]-[d(GC(ps)T(ps)ACG)] (ps, phosphorothioate; PS-DNA), with their complementary RNA have been investigated by combined use of (1)H NMR and restrained molecular dynamics calculation. Compared to those obtained for the unmodified duplex structures (PO-DNA.RNA), the NOE cross-peak intensities are virtually identical for the PS-DNA.RNA hybrid duplexes. The structural analysis on the basis of the NOE restraints reveals that all of the three DNA.RNA duplexes take a A-form conformation and that there is no significant difference in the base stacking for the DNA.RNA hybrid duplexes. On the other hand, the NOE cross-peak intensities of the protons around the central T(ps)A step of the PS-DNA.DNA duplexes are apparently different from those of PO-DNA. DNA. The chemical shifts of H8/6 and H1' at the T(ps)A step are also largely different among PS-DNA.DNAs and PO-DNA.DNA, suggesting that the DNA.DNA structure is readily changed by the introduction of the phosphorothioate groups to the central T(p)A step. The structure calculations indicate that all of these DNA.DNA duplexes are B-form although there exist some small differences in helical parameters between the [R(p),R(p)]- and [S(p),S(p)]PS-DNA.DNA duplexes. The melting temperatures (T(m)) were determined for all of the duplexes by plotting the chemical shift change of isolated peaks as a function of temperature. For the PS-DNA.RNA hybrid duplexes, the [S(p),S(p)] isomer is less stable than the [R(p),R(p)] isomer while this trend is reversed for the PS-DNA.DNA duplexes. Consequently, although the PS-DNA.RNA duplexes take the similar A-form structure, the duplex stability is different between PS-DNA.RNA duplexes. The stability of the DNA.RNA duplexes may not be governed by the A-form structure itself but by some other factors such as the hydration around the phosphorothioate backbone, although the T(m) difference of the DNA.DNA duplexes could be explained by the structural factor.

Mutagenesis by an Antisense Oligonucleotide and Its Degradation Product

“…The sulphur within the phosphodiester backbone is likely to be recognised as a lesion by the nucleotide excision repair (NER) pathway ultimately resulting in the excision of an encompassing fragment of 27-29nt in length (Huang et al, 1992, Svoboda et al, 1993. In support of this, the introduction of phosphorothioate bonds in a DNA double helix has been reported to cause structural alterations (Kanaori et al, 1999).…”

Gene Silencing in Mammalian Cells with Light‐Activated Antisense Agents

“…The noncaged PS DNA/RNA hybrid melted at A C H T U N G T R E N N U N G approximately 39 8C, which agreed with that of similar PS DNAs. [22] Irradiation at 365 nm for 5 min completely restored hybridization for both caged antisense agents (Table 1).…”

Theoretical Study on Steric Effects of DNA Phosphorothioation: B-Helical Destabilization in Rp-Phosphorothioated DNA

“…Our calculations show that steric hindrance of neighboring pentose could lead to significant repulsion to the sulfur atom in the case of B-helical Rp-DNA, while no such effect was observed in the case of Spconfiguration. 32 …”