2D¹H and³¹P NMR Spectra and Distorted A-DNA-Like Duplex Structure of a Phosphorodithioate Oligonucleotide

Abstract

Assignment of the 1H and 31P NMR spectra of a phosphorodithioate modified oligonucleotide decamer duplex, d(CGCTTpS2-AAGCG)2 (10-mer-S; a site of dithioate substitution is designated with the symbols pS2-), was achieved by two-dimensional homonuclear TOCSY, NOESY and 1H-31P Pure Absorption phase Constant time (PAC) heteronuclear correlation spectroscopy. In contrast to the parent palindromic decamer sequence (1) which has been shown to exist entirely in the duplex B-DNA conformation under comparable conditions (100 mM KCl), the dithiophosphate analogue forms a hairpin loop. However, the duplex form of the dithioate oligonucleotide can be stabilized at lower temperatures, higher salt and strand concentration. The solution structure of the decamer duplex was calculated by an iterative hybrid relaxation matrix method (MORASS) combined with 2D NOESY-distance restrained molecular dynamics. These backbone modified compounds, potentially attractive antisense oligonucleotide agents, are often assumed to possess similar structure as the parent nucleic acid complex. Importantly, the refined structure of the phosphorodithioate duplex shows a significant deviation from the parent unmodified, phosphoryl duplex. An overall bend and unwinding in the phosphorodithioate duplex is observed. The structural distortion of the phosphorodithioate duplex was confirmed by comparison of helicoidal parameters and groove dimensions. Especially, the helical twists of the phosphorodithioate decamer deviate significantly from the parent phosphoryl decamer. The minor groove width of phosphorodithioate duplex 10-mer-S varies between 8.4 and 13.3 A which is much wider than those of the parent phosphoryl decamer d(CGCTTAAGCG)2 (4.2 approximately 9.4 A). The larger minor groove width of 10-mer-S duplex contributes to the unwinding of the backbone and indicates that the duplex has an overall A-DNA-like conformation in the region surrounding the dithiophosphate modification.

Phosphorothioate oligonucleotides reduce mitochondrial outer membrane permeability to ADP

“…This is critical because the binding energy is the difference between the energy of interaction between the protein and the oligonucleotide and the energies of dehydration of the interacting surfaces. The low-charge density of the sulfur also reduces the enthalpy needed to strip small ions before binding to VDAC (6). This property also increases the polarizability (37) of the sulfur atoms, strengthening the interaction with lower charge density groups found in proteins.…”

NMR Solution Structure of the N3‘ → P5‘ Phosphoramidate Duplex d(CGCGAATTCGCG)2 by the Iterative Relaxation Matrix Approach

“…Such RNAs have a structural preference for A-form helical regions with sugars in a C3′endo conformation (6). First generation antisense oligonucleotides such as phosphorothioates and methylphosphonates, based on deoxyribose sugars, exist predominantly in C2′-endo conformations and prefer a B-form helix typical of DNA (7)(8)(9)(10)(11). These first generation agents generally form complexes with target RNAs that are less stable than similar complexes with unmodified DNA (7,8,(12)(13)(14).…”

Structure and Dynamics of a DNA.cntdot.RNA Hybrid Duplex with a Chiral Phosphorothioate Moiety: NMR and Molecular Dynamics with Conventional and Time-Averaged Restraints

“…The few structural studies in modified oligonucleotides carried out so far focus on DNAONA duplexes (Heinemann et al, 1991;Gao et al, 1992;Stolarski et al, 1992). In most cases, the modification appears to fit well in the general B-type structure of the duplex, but in the case of phosphorodithioates significant distortions from the B-family of structures have been detected (Cho et al, 1993). Although mRNA is the most attractive target for the antisense chemotherapeutical strategy, virtually nothing is known about the structure of oligonucleotide analogs in DNA'RNA duplexes.…”