Unusual conformation of a 3?-thioformacetal linkage in a DNA duplex

Abstract

The DNA.DNA duplex d(CGCGTTSCH2OTTGCGC).d(GCGCAAAACGCG) (designated duplex III) containing a 3'-thioformacetal (3'-TFMA) linkage in the center of the sequence was characterized in detail by two- and three-dimensional homonuclear NMR spectroscopy. The NMR results were analyzed and compared with those of two duplexes of the same sequence: One is an unmodified reference sequence and the other contains a formacetal (OCH2O) linkage at the central T--T step (designated duplex I and duplex II, respectively). In general, the NMR spectra of duplex III closely resemble those of the analogous duplexes I and II, suggesting an overall B-type structure adopted by the 3'-TFMA-modified duplex III. Nonetheless, the detection of several distinct spectral features originating from the protons at the T6(3'-SCH2O)T7 modification site is indicative of a local conformation that is clearly different from the corresponding region in duplexes I and II. The 3'-thioformacetal linker, in contrast to the formacetal (FMA) linkage, cannot be accommodated in a conformation usually found in natural nucleic acid duplexes. As a consequence, the 3'-TFMA-modified T6 sugar adopts an O4'-endo form (an intermediate structure between the usual C2'-endo and C3'-endo forms). This change is accompanied by a change in the epsilon (C4'-C3'-S3'-CH2) dihedral angle and by subsequent adjustments of other torsion angles along the backbone. Notably, this conformational readjustment at the T6-T7 backbone linkage is localized; its collective result has negligible effect on base-base stacking of the T6 and T7 residues. A close examination of the COSY data in all three duplexes reveals a subtle variation in sugar geometry, with more S-type character adopted by the modified duplexes II and III. The results of this study illustrate that, although the difference between FMA and 3'-TFMA linkages is merely in the substitution of the T6(O3') in the former by a sulfur atom in the latter, the stereoelectronic difference in a single atom can induce significant local structural distortion in an otherwise well-structured oligonucleotide duplex.

An approach to the structure determination of nucleic acid analogues hybridized to RNA. NMR studies of a duplex between 2′-OMe RNA and an oligonucleotide containing a single amide backbone modification

“…Region B and C ( Figure 5) are selected to start the sequential assignment. Region B is attractive to start the assignment of the deoxyribose strand, because no signals of the 2'-OMe RNA -:~~~~~~~~Q 4.i "-' 0i 8.0 6.0 4 0 2.0 Dl (ppm) Figure 5.…”

A model for DNA polymerase translocation: worm‐like movement of DNA within the binding cleft

“…This mechanism can explain our earlier findings that the Vm~x values of DNA polymerization are sufficiently higher for many polymerases if ribooligonucleotides (or deoxyribooligonucleotides with the 3'-terminal ribonucleotide) have been used as a primer [31]. The A-like form is preferable for RNA-DNA hybrid duplexes [32,33] (and for some short duplexes which contains the ribonucleotide insertion [34]), hence the reverse B ~ A transition within the binding channel should have a lower energetic barrier in these cases. Mikita and Beardsley [35,36] have shown that DNA replication can be arrested by structural lesion of arabinosylcytosine in.…”

“…The A-like form is preferable for RNA-DNA hybrid duplexes [32,33] (and for some short duplexes which contains the ribonucleotide insertion [34]), hence the reverse B ~ A transition within the binding channel should have a lower energetic barrier in these cases. Mikita and Beardsley [35,36] have shown that DNA replication can be arrested by structural lesion of arabinosylcytosine in.…”

Homonuclear Three-Dimensional NMR of Biomolecules

“…Such methods have been applied for the analysis of the NOEs of a 12-mer DNA duplex containing a GG mismatch 99 and for analysis of a modified 12-mer DNA duplex. 100 In both cases the 3D spectra clarified unusual NOEs, with important consequences for the geometry.…”

“…It has been demonstrated that 3D NOESY-NOESY spectra of good quality can be recorded for a 22-mer duplex. 11 Similarly, 3D NOESY-NOESY has been applied to a 40-base threestranded DNA junction, 101 where it allowed the completion of the proton assignments and confirmed the unusual geometry at the junction, and to a hybrid DNA-RNA dodecamer duplex, 102 where it resolved superimposed cross peaks of the RNA strand.…”