Effects of the Introduction of L-Nucleotides into DNA. Solution Structure of the Heterochiral Duplex d(G-C-G-(L)T-G-C-G).cntdot.d(C-G-C-A-C-G-C) Studied by NMR Spectroscopy
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Abstract
The effect of the substitution of a L-nucleoside for a D-nucleoside in the duplex
d(G-C-G-T-G-C-G).d(C-G-C-A-C-G-C) was studied by UV and NMR spectroscopy. These unnatural
oligonucleotides have potential for antisense DNA technology [Damha, M. J., Giannaris,
P. A., & Marfey, P. (1994) Biochemistry (preceding paper in this issue)]. The thermal
stability of such duplexes is lower than that of the natural one and is dependent
on the nucleotide type and/or sequence. Interestingly, inversion of the chirality
of thymidine but not adenosine coincides with a large stabilizing enthalpy change.
The structure of the heterochiral duplex d(G1-C2-G3-(L)T4-G5-C6-G7).d(C8-G9-C10-A11-C12-G13-
C14), where (L)T denotes the mirror image of the natural thymidine, has been determined
by NMR spectroscopy. The sugar conformation was determined using the sum of coupling
constants and the distances using a model free relaxation matrix approach. The torsion
angles of the backbone follow from 3JHH, 3JHP, and 4JHP coupling constants. The structure
of the duplex was calculated by metric matrix distance geometry followed by simulated
annealing. The structure is close to that of B-DNA. The base pair formed by (L)T and
A is of the Watson-Crick type. All sugars adopt an S-type pucker. The incorporation
of the L-sugar in the duplex is accomplished by changes in the backbone torsion angles
around the phosphates and the glycosidic torsion angle of (L)T. The modification induces
changes in the natural strand as well. The structure exhibits an unusual interaction
between the aromatic rings of the (L)T4.A11 and G3.C12 base pairs, which provides
a plausible explanation of the unusual thermodynamic properties of the duplex.