Position-dependent effects on stability in tricyclo-DNA modified oligonucleotide duplexes

Abstract

A series of oligodeoxyribonucleotides and oligoribonucleotides containing single and multiple tricyclo(tc)-nucleosides in various arrangements were prepared and the thermal and thermodynamic transition profiles of duplexes with complementary DNA and RNA evaluated. Tc-residues aligned in a non-continuous fashion in an RNA strand significantly decrease affinity to complementary RNA and DNA, mostly as a consequence of a loss of pairing enthalpy ΔH. Arranging the tc-residues in a continuous fashion rescues T(m) and leads to higher DNA and RNA affinity. Substitution of oligodeoxyribonucleotides in the same way causes much less differences in T(m) when paired to complementary DNA and leads to substantial increases in T(m) when paired to complementary RNA. CD-spectroscopic investigations in combination with molecular dynamics simulations of duplexes with single modifications show that tc-residues in the RNA backbone distinctly influence the conformation of the neighboring nucleotides forcing them into higher energy conformations, while tc-residues in the DNA backbone seem to have negligible influence on the nearest neighbor conformations. These results rationalize the observed affinity differences and are of relevance for the design of tc-DNA containing oligonucleotides for applications in antisense or RNAi therapy.

Figures

Figure 1.

Chemical structure of tricyclo(tc)-DNA.

Figure 2.

Graphical representation of the thermodynamic data of duplex formation (standard conditions, 25°C, kcal/mol) from van’t Hoff analysis of the UV-melting curves of oligonucleotides 1, 7, 9 and 11 in duplex with either complementary DNA (a) or RNA (b).

Figure 3.

CD spectra of oligonucleotides 1, 3, 7 and 9 in complex with complementary DNA (a) and RNA (b). Experimental conditions as for UV-melting curves, T = 25°C.

Figure 4.

Graphical representation of the thermodynamic data of duplex formation (standard conditions, 25°C, kcal/mol) from van’t Hoff analysis of the UV-melting curves of oligonucleotides 12, 18, 20 and 11 in duplex with either complementary DNA (a) or RNA (b).

Figure 5.

CD spectra of oligoribonucleotides 12, 14, 18 and 20 in complex with complementary DNA (a) and RNA (b). Experimental conditions as for UV-melting curves, T = 25°C.

Figure 6.

Strereoviews of the central section of the duplexes of (a) 3/DNA, (b) 3/RNA, (c) 14/DNA and (d) 14/RNA, containing the modified tc-DNA- as well as its 3′- and 5′-flanking natural base pairs after a total of 400 ps dynamic simulation followed by energy minimization using the AMBER force field as implemented in the software package Hyperchem™.