Acyclic, achiral enamide nucleoside analogues. The importance of the C=C bond in the analogue for its ability to mimic natural nucleosides

Article abstract of DOI:10.1039/b307394g

The conformations of an acyclic, achiral enamide thymidine analogue 1 have been studied by model building and geometry calculations, as well as by NMR NOE and UV experiments. The results indicate that there are no significant barriers to rotation around any of the σ bonds, in particular the N1-C1 ′ enamide bond, and that the analogue should be able to accommodate conformations that mimic the conformations of natural nucleosides in A- and B-type helices quite well. For comparison the saturated analogue 2 has been prepared and built into oligonucleotides. It is shown that incorporation of 2 in oligonucleotides results in a much larger depression of the melting temperature (ΔT _m - 10 to -12.5 °C) than does incorporation of 1 (ΔT _m - 5 to -6.5 °C).

Full text of DOI:10.1039/b307394g

View Article Online / Journal Homepage / Table of Contents for this issue
Acyclic, achiral enamide nucleoside analogues. The importance of
the C᎐C bond in the analogue for its ability to mimic natural
᎐
nucleosides
Asger B. Petersen, Michael Å. Petersen, Ulla Henriksen, Steen Hammerum and Otto Dahl*
Department of Chemistry, University of Copenhagen, The H. C. Ørsted Institute,
Universitetsparken 5, DK-2100 Copenhagen, Denmark. E-mail: dahlo@kiku.dk;
Fax: ϩ45 3532 0212; Tel: ϩ45 3532 0176
Received 30th June 2003, Accepted 15th August 2003
First published as an Advance Article on the web 4th September 2003
The conformations of an acyclic, achiral enamide thymidine analogue 1 have been studied by model building and
geometry calculations, as well as by NMR NOE and UV experiments. The results indicate that there are no
significant barriers to rotation around any of the σ bonds, in particular the N1–C1Ј enamide bond, and that the
analogue should be able to accommodate conformations that mimic the conformations of natural nucleosides in
A- and B-type helices quite well. For comparison the saturated analogue 2 has been prepared and built into
oligonucleotides. It is shown that incorporation of 2 in oligonucleotides results in a much larger depression of the
melting temperature (∆Tm Ϫ10 to Ϫ12.5 ЊC) than does incorporation of 1 (∆Tm Ϫ5 to Ϫ6.5 ЊC).
structure of 1 high enough to result in a substantial barrier to
Introduction
rotation around the N1–C1Ј bond? We have also prepared the
hitherto unknown6 saturated analogue 2 and show here that
oligonucleotides containing this analogue bind much less well
to DNA and RNA than the oligonucleotides containing 1 in the
same positions.
Nucleoside analogues are potential antiviral and anticancer
agents,1 and may improve the in vivo properties of antisense
oligonucleotides which are promising highly selective drugs.2
Many acyclic nucleoside analogues have shown high antiviral
activity, but are usually poor monomer building blocks for anti-
sense-directed purposes, because of their flexibility which
results in reduced binding of the modified antisense oligo-
nucleotide to a target mRNA. Thus, a reduction of 6–13 ЊC in
the melting temperature (Tm, the temperature at which half of
a duplex is dissociated) of DNA–RNA 17-mer duplexes has
been found when the DNA oligonucleotide is modified with one
acyclic nucleoside analogue in the middle of the sequence.3
We have prepared4 and built into oligodeoxyribonucleotides5
an acyclic, achiral enamide nucleoside analogue, 1-[3-hydroxy-
2-(hydroxymethyl)prop-1-enyl]thymine 1 (Chart 1). The ana-
logue was designed to mimic natural nucleosides, i.e. to be able
to exist in conformations which place the two hydroxy groups
and thymine close to the positions of the 3Ј-hydroxy group, the
5Ј-hydroxy group, and thymine of thymidine located in both
natural dsDNA (B-type double helices) and dsRNA (A-type
double helices). The analogue was arrived at by model building,
Results and discussion
Model building
At first sight 1 seems far removed from a normal thymine
nucleoside structure. In order to illustrate that 1 may be able to
mimic a thymine nucleoside in A- and B-type double helices the
structure of 1 was drawn with ChemDraw and PM3 minimized
with Hyperchem 7.7 Overlay pictures of the resulting structure
1 with dT or 2Ј-F-araT structures, taken from published crystal
structures of double helices containing dT in a B-helix,8 dT in
an A-helix,9 and 2Ј-F-araT in a modified B-helix,10 are shown
in Fig. 1. In these overlay pictures, only rotations around the
N1–C1Ј, C2Ј–C3Ј, and C2Ј–C4Ј bonds (dihedral angles χ, δ,
and γ, respectively) of 1 were allowed, all bond lengths and
other angles were unchanged. The overlay pictures were arrived
at by distance minimization, performed with the computer
program Macromodel 7,11 between the same atoms of the bases
(N1, O2, N3, and O4), and between O3Ј in 1 and O3Ј in the
sugars, and O4Ј in 1 and O5Ј in the sugars. No energy minimiza-
tions were performed.
and the C᎐C double bond seemed important to obtain good
᎐
overlap between the above-mentioned essential groups. How-
ever, oligonucleotides containing one or two 1 were found to
bind to complementary DNA and RNA strands with a reduced
Tm (2–6.5 ЊC per single introduced base modification).5 The
present paper describes our studies on the preferred con-
formations of 1 by NMR, UV, and geometry calculations. We
address the following questions: Are conformations of 1 which
fit into A- or B-type helices minima, or how much energy is
required to reach these conformations from the minimum
energy conformation(s)? Is conjugation in the enamide
Chart 1 Dihedral angles χ = C2Ј–C1Ј–N1–C6; γ = C1Ј–C2Ј–C4Ј–O4Ј;
δ = C1Ј–C2Ј–C3Ј–O3Ј.
Fig. 1
T h i s j o u r n a l i s © T h e R o y a l S o c i e t y o f C h e m i s t r y 2 0 0 3
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 3 2 9 3 – 3 2 9 6
3293

View Article Online
Table 1 Distances between similar atoms in the overlayed structures shown in Fig. 1 and selected dihedral angles as defined for structure 1
Distances/Å
Dihedral angles/Њ
Structure
O3Ј–O3Ј
O4Ј–O5Ј
N1–N1
O2–O2
N3–N3
O4–O4
χ
δ
γ
A
B
C
0.259
0.375
0.275
0.098
0.288
0.089
0.219
0.310
0.404
0.056
0.087
0.170
0.079
0.157
0.082
0.130
0.293
0.297
Ϫ8.4
Ϫ2.8
Ϫ13.0
51.8
64.5
114.6
136.8
176.8
114.0
The minimized distances between analogous atoms in the
superimposed structures and the dihedral angles χ, δ, and γ are
given in Table 1. None of these distances are found to be larger
than 0.4 Å, and the dihedral angle χ is between Ϫ3 and Ϫ13Њ.
These models suggest that 1 should be able exist in conform-
ations that mimic the conformations of natural nucleosides
in A- and B-type helices quite well. Estimates of the energies
necessary to reach these conformations from the minimum
energy conformation of 1 are given in the following section.
of the 4Ј-OH signal, and 7% of the CH3 thymine signal. The
similar NOE’s of the H1Ј and the H4Ј signals corroborate that
a population of conformers exists in which χ is substantial, as
found by the above calculations; a significant population of
nearly planar conformations (χ = 0) should give a much larger
NOE of the H4Ј signals. However, some conjugation is indi-
cated by UV spectroscopy. Increasing the extent of conjugation
in a system results in a bathochromic shift and an increase in
intensity of the absorption. The UV spectrum of 1 in MeOH
compared with those of some saturated alkyl analogues shows
a small bathochromic shift and a small increase in intensity.
Thus λmax for the longer-wavelength band of 1 is 273 nm (ε =
1.05ؒ104 l molϪ1 cmϪ1), compared to 269 nm (ε = 9.55ؒ103
l molϪ1 cmϪ1) for the saturated analogue 1-(2,3-dihydroxy-2-
hydroxymethylpropyl)thymine and 270 nm (ε = 9.50ؒ103 l molϪ1
cmϪ1) for the saturated analogue 2.
DFT calculations
Density functional theory (DFT) and ab initio calculations were
used to find the minimum energy conformations of 1 and to
estimate the energy barriers to rotation around the N1–C1Ј
bond (dihedral angle χ). B3LYP/6-31G(d) calculations were
used to determine the minimum-energy structure as well as the
geometry of other stationary points and to obtain zero-point
vibrational energy contributions, and the MP2/6-31G(d)
method was used for energy calculations at these stationary
points. The calculations were performed with the Gaussian98
suite of programs.12
The dihedral angle between the ring and the exocyclic double
bond (χ) is 39Њ in the minimum energy conformation. Rotation
around the N1–C1Ј bond (i.e., variation of χ) involves an
energy barrier of 10 kJ molϪ1 (MP2-calculations); the highest
energy conformation is nearly planar (χ = 4Њ). In all calculated
conformations one of the OH groups forms a hydrogen bond
to the other OH group, as expected for a gas phase species; this
precludes an estimate of barriers to rotation around the
C2Ј–C3Ј and C2Ј–C4Ј bonds. Calculations on a similar struc-
ture without the hydroxy groups [1-(2Ј-methylprop-1-enyl)u-
racil] gave no indications of any substantial barrier to rotation
around these bonds, and showed properties similar to 1 (χ = 52Њ
in the minimum conformation, barrier 21 kJ molϪ1 (MP2) or
14 kJ molϪ1 (B3LYP) for the highest energy conformation in
which χ is 0Њ).
From these solution experiments we conclude that conform-
ations with a substantial deviation from χ = 0Њ are significantly
populated, although the UV results indicate that some π-orbital
overlap is present.
Preparation of the saturated analogue 2 and binding studies of
oligonucleotides containing 2
The saturated analogue 2 was prepared from the known alcohol
314(Scheme 1). Thus, 3-benzoylthymine15 was coupled with 3
under Mitsunobu conditions to give 4, which was hydrolysed
to 2 in a good yield. Alkylation of 3-benzoylthymine at N-1
was confirmed by NOE. The analogue 2 was mono-di-
methoxytritylated to give 5 (racemic) in a fair yield using
one equivalent of dimethoxytrityl chloride and then 5 was
converted to the phosphoramidite 6 in a standard way.
These results strongly suggest that there are no significant
barriers to rotation around any of the σ bonds in 1, in particu-
lar around the N1–C1Ј enamide bond, which was originally
expected to have a substantial barrier to rotation due to con-
jugation between the π system of T and the C1Ј–C2Ј double
bond. Any conjugative stabilization is apparently outweighed
by other factors which results in a minimum energy conform-
ation with χ far from 0. A similar result was obtained in an X-ray
crystal structure of an enamide (N-[2-methyl-1-(2-naphthyl)-
prop-1-enyl]acetamide) where the enamide dihedral angle was
found to 78Њ.13 We have so far been unable to obtain crystalline
1 suitable for X-ray crystallography.
Scheme 1 a) Ph3P, PriOOCN᎐NCOOPri, THF, rt overnight; b) 1 M
aq. NaOH–dioxane, rt overnight; c) 80% acetic acid, 50 ЊC, 2 h; d)
DMTCl, pyridine, rt overnight; e) NCCH2CH2OP(NPri2)2, tetrazole,
CH3CN, rt.
᎐
Two oligodeoxyribonucleotides containing the saturated
analogue 2 (TS) were prepared by standard phosphoramidite
solid phase oligonucleotide synthesis and their hybridization
properties compared with those containing the unsaturated
analogue 1 (TU) in the same positions (Table 2). It is seen that
the incorporation of TS gives a much larger depression of
the melting temperature (∆Tm Ϫ10.0 to Ϫ12.5 ЊC) against
DNA and RNA complements than TU (∆Tm Ϫ5.0 to Ϫ6.5 ЊC).
Although the TS unit is racemic, the melting curves had similar
shapes (similar hyperchromicity and wideness of the transition)
NMR (NOE) and UV experiments
Information on the preferred conformation(s) of 1 in solution
can be obtained from 1H NMR NOE (Nuclear Overhauser
Enhancements) and UV absorption experiments. For 1 in a
nearly planar conformation there should be a large NOE
between H6 and H4Ј(χ = 0Њ) or H6 and H1Ј(χ = 180Њ), and the
UV absorption band should move towards longer wavelengths
due to conjugation. Irradiation at H6 of 1 (in DMSO-d6) gave
a NOE of 4.5% of the H4Ј signals, 3% of the H1Ј signal, 2%
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 3 2 9 3 – 3 2 9 6
3294

View Article Online
Table 2 Hybridization data (Tm, ЊC) for modified and unmodified oligonucleotides with DNA and RNA complementsa
b
b
dA14
∆Tm
rA14
∆Tm
dT14
36.0
31.0
26.0
33.5
28.0
23.5
dT7TUT6
dT7TST6
Ϫ5.0
Ϫ10.0
Ϫ5.5
Ϫ10.0
b
b
dGTGAGATGC
∆Tm
rGTGAGATGC
∆Tm
dGCATCTCAC
39.0
28.0
14.0
41.0
27.5
18.0
dGCATUCTUCAC
dGCATSCTSCAC
Ϫ5.5
Ϫ12.5
Ϫ6.5
Ϫ11.5
a Tm was determined by measuring absorbance at 260 nm against increasing temperature (0.5 ЊC steps) on equimolar mixtures (3 µM in each strand)
of modified oligomer and its complementary DNA or RNA strand in medium salt buffer (10 mM Na2HPO4, 100 mM NaCl, 0.1 mM EDTA, pH
7.0). TU and TS are explained in the text. b Change in Tm per modification.
1
for unmodified T, TU, and TS, which indicate that both stereo-
isomers of TS bind with similar low affinity to their
complements.
for H): δH 8.29 (1H, br s, NH), 7.07 (1H, q, J 1.2, H-6), 3.94
(2H, d, J 7.9, CH2N), 3.86 (4H, ABX system, ∆ 127 Hz,
JAB 12.6, Jax = JBX 3.1, CH2O), 2.03 (1H, m, CH), 1.92 (3H, d,
J 1.2, CH3T), 1.47 (3H, s, CH3), 1.43 (3H, s, CH3). δC 164.4,
151.1, 141.4, 110.3, 98.6, 61.0, 48.4, 32.8, 27.8, 20.0, 12.2. FABϪ
MS: 253.2 (M Ϫ Hϩ), calc. 253.1. Found: C, 56.6; H, 7.1;
N, 11.25. Calc. for C12H18N2O4: C, 56.7, H, 7.1, N, 11.0%.
Conclusion
The analogue 1 was designed to be conformationally restricted
by a C᎐C bond in order to promote preorganization to mimic
᎐
natural nucleosides. The modeling shown in Fig. 1 indicates
that this is indeed successful. DFT and ab initio calculations
indicate that there is no significant energy barrier to rotation
around the σ bonds necessary to attain the modelled conform-
ations. These calculations describe the properties of gas phase
species, but NMR NOE and UV experiments in solution
(DMSO-d6 and MeOH, respectively) indicate that conform-
ations which are rotated around the N1–C1Ј bond (χ deviating
from 0Њ) are substantially populated. The importance of the
1-[3-Hydroxy-2-(hydroxymethyl)prop-1-yl]thymine (2)
Compound 4 (0.940 g, 3.70 mmol) was dissolved in 80% aq.
acetic acid (50 ml) and stirred at 50 ЊC for 2 h. The reaction
mixture was evaporated in vacuo and the solid residue recrystal-
lised from methanol to give 2 (0.596 g, 75%) as colourless crys-
tals, mp 146–147 ЊC (lit.6 mp 133–134 ЊC). NMR (DMSO-d6):
δH 11.2 (1H, br s, NH), 7.40 (1H, s, H-6), 4.52 (2H, br s, OH),
3.60 (2H, d, J 7.0, CH2N), 3.37 (4H, d, J 5.3, CH2O), 1.89 (1H,
m, CH), 1.73 (3H, s, CH3). δC 164.2, 151.1, 142.0, 108.1, 59.1,
46.8, 42.8, 11.9. FABϪ MS: 212.8 (M Ϫ Hϩ), calc. 213.1. Found:
C, 50.5; H, 6.6; N, 12.85. Calc. for C9H14N2O4: C, 50.5; H, 6.6;
N, 13.1%.
C᎐C bond for the ability of 1 to mimic a natural dT unit in
᎐
oligonucleotides is shown by comparing the melting tem-
peratures (Tm) of oligonucleotides, modified with 1 or the
saturated analogue 2, when hybridized to their DNA and RNA
complements (Table 2). Clearly 1 mimics a dT unit much better
than 2, although the affinity of both analogues is lower than
that of natural dT.
1-[3-(Dimethoxytrityloxy)-2-(hydroxymethyl)prop-1-yl]thymine
(5)
The results presented here indicate that nothing in the
structure of 1 should prevent good binding of oligonucleotides
modified with 1 to both DNA and RNA complements. Never-
theless 1 is a poor substitute for natural dT. Possible reasons for
this could be a disruption of solvation at the rather unpolar 1,
or subtle changes in the geometry around the modification.
An answer to these questions must await an X-ray structure
determination of a modified duplex.
Compound 2 (0.410 g, 1.92 mmol) was coevaporated with dry
pyridine and dissolved under nitrogen in dry pyridine (10 ml).
Dimethoxytrityl chloride (0.65 g, 1.92 mmol) was added and
the mixture stirred at rt in the dark overnight. Pyridine was
removed in vacuo and the residue purified by flash chromato-
graphy on silica (Merck silica 60, 0.040–0.063 mm, eluted with
ethyl acetate–heptane–triethylamine 89 : 10 : 1 v/v/v) to give 5
(0.354 g, 36%) as a colourless foam. NMR (CDCl3, 300 MHz
for 1H): δH 8.3 (1H, br s, NH), 7.39 (2H, d, J 7.3, Ar), 7.32–7.23
(7H, m, Ar), 6.96 (1H, q, J 1.2, H-6), 6.83 (4H, d, J 8.8, Ar),
3.95 (2H, ABX system, ∆ 21.1 Hz, JAB 14.1, JAX 4.7, JBX 6.0,
CH2N), 3.79 (6H, s, CH3O), 3.50 (2H, ABX system, ∆ 12.8 Hz,
JAB 12.1, JAX 6.6, JBX 4.9, CH2OH), 3.14 (2H, ABX system,
∆ 101.9 Hz, JAB 9.7, JAX 4.8, JBX 7.4, CH2ODMT), 2.19 (1H,
m, CH), 1.79 (3H, d, J 1.2, CH3T). δC 164.3, 158.3, 151.9,
144.4, 141.2, 135.5, 135.4, 129.7, 127.7, 127.6, 126.7, 112.9,
110.4, 86.3, 61.5, 60.3, 55.0, 46.5, 41.6, 12.1. FABϩ MS: 517.0
(M ϩ Hϩ), calc. 517.2.
Experimental
1-[(2,2-Dimethyl-1,3-dioxan-5-yl)methyl]thymine (4)
Triphenylphosphine (2.67 g, 10.2 mmol), 3-benzoylthymine15
(2.32 g, 10.1 mmol), and 2,2-dimethyl-5-(hydroxymethyl)-1,3-
dioxane14 (1.34 g, 9.2 mmol) were coevaporated with
acetonitrile and dissolved under nitrogen in dry THF (50 ml).
Diisopropyl azodicarboxylate (DIAD) (2.34 g, 11.6 mmol) in
dry THF (20 ml) was added, and the mixture stirred overnight
at rt. Following evaporation of solvents in vacuo the residue was
dissolved in 1 M aq. NaOH–dioxane (1 : 1 v/v, 40 ml) and the
solution stirred overnight at rt. The basic solution was extracted
with diethyl ether (3 × 30 ml), the ether phase extracted with
0.01 M aq. NaOH, the combined aq. phases neutralised to
pH ca. 8 with acetic acid, and the product extracted with di-
chloromethane (7 × 30 ml). The dichloromethane solution was
dried (Na2SO4), the solvent removed in vacuo, and the yellow
crude oil crystallised from acetonitrile to give 4 (1.39 g, 60%) as
colourless crystals, mp 171.5–173 ЊC. NMR (CDCl3, 300 MHz
1-[3-(Dimethoxytrityloxy)-2-[2-cyanoethoxy(diisopropylamino)-
phosphinoxymethyl]prop-1-yl]thymine (6)
To 5 (0.187 g, 0.362 mmol) in dry acetonitrile (2 ml) under
N2 was added 2-cyanoethyl N,N,NЈ,NЈ-tetraisopropylphos-
phorodiamidite (0.230 g, 0.76 mmol), followed by tetrazole
(0.025 g, 0.36 mmol). After stirring for 30 min the solvent was
removed in vacuo, the residue dissolved in dichloromethane
(6 ml), washed with sat. aqueous NaHCO3 (2 ml), dried
(Na2SO4), and the dichloromethane removed in vacuo. The
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 3 2 9 3 – 3 2 9 6
3295

View Article Online
2 Oligonucleotides as Therapeutic Agents, ed. D. J. Chadwick and
G. Cardew, Ciba Foundation, John Wiley & Sons Ltd, Chichester,
England, 1997; Antisense Research and Application, ed. S. T. Crooke,
Springer Verlag, Berlin, 1998; E. Uhlmann, Curr. Opin. Drug
Discovery Dev., 2000, 3, 203.
3 P. Nielsen, L. H. Dreiøe and J. Wengel, Bioorg. Med. Chem., 1995,
3, 19.
4 D. S. Pedersen, T. Boesen, A. B. Eldrup, B. Kiær, C. Madsen,
U. Henriksen and O. Dahl, J. Chem. Soc., Perkin Trans. 1, 2001,
1656.
5 T. Boesen, D. S. Pedersen, B. M. Nielsen, A. G. Petersen,
U. Henriksen, B. M. Dahl and O. Dahl, Bioorg. Med. Chem. Lett.,
2003, 13, 847.
residue was purified by flash chromatography on silica (Merck
silica 60, 0.040–0.063 mm, eluted with ethyl acetate–heptane–
triethylamine 60 : 39 : 1 v/v/v) and the product precipitated in
pentane to give 6 (0.077 g, 30%) as a colourless foam. 1H NMR
(CDCl3): δH 8.2 (1H, br s, NH), 7.40–7.20 (9H, m, Ar), 6.93
and 6.91 (1H, s, H-6, 2 diastereomers), 6.82 (4H, d, J 8.5, Ar),
3.79 (6H, s, CH3O), 3.75–3.24 (10H, m, 3 × CH2O, CH2N, 2 ×
CHN), 2.58 and 2.54 (2H, t, J 6.3, CH2CN, 2 diastereomers),
2.43 and 2.36 (1H, m, CH(CH3)2, 2 diastereomers), 1.78 (3H,
s, CH3T), 1.17 (6H, d, J 6.7, CH(CH3)2), 1.11 (6H, d, J 6.7,
CH(CH3)2). δC 164.3, 158.7, 151.0, 144.9, 141.8, 141.6, 136.1,
130.1, 128.2, 128.0, 127.0, 117.8, 113.3, 110.1, 86.4, 62.7,
62.5, 62.3, 62.1, 61.8, 61.7, 58.7, 58.5, 55.4, 48.4, 48.2, 43.5,
43.2, 40.6, 40.5, 24.9, 24.8, 20.7, 12.4. δP 149.1, 149.0. FABϪ
MS: 716.1 (M Ϫ Hϩ), calc. 715.3.
6 After the completion of our work 2 has been prepared by another
route: S. Guillarme, S. Legoupy, A.-M. Aubertin, C. Olicard,
N. Bourgougnon and F. Huet, Tetrahedron, 2003, 59, 2177.
7 ChemDraw Ultra version 6.01, www.cambridgesoft.com;
HyperChem7, www.hyper.com.
8 C. L. Kielkopf, S. Ding, P. Kuhn and D. C. Rees, J. Mol. Biol., 2000,
296, 787 (Nucleic Acid Database BD0023).
9 Y. Xiong and M. Sundaralingam, Nucleic Acids Res., 2000, 28, 2171
(Nucleic Acid Database AH0005).
Oligonucleotide syntheses. Oligonucleotide syntheses were
performed in 0.2 µmol scale on a Biosearch 8750 DNA
Synthesizer using standard conditions with unmodified
phosphoramidites (Cruachem), or 6 and tetrazole activation.
The modified phosphoramidite 6 (0.05 M in acetonitrile) was
manually coupled for 6 min with a DMT efficiency of 98–99%.
The oligonucleotides (precipitated by ethanol), dT7TST6, >95%
pure (CE), M 4167.7 (ESI-MS, calc. 4167.7), and dGCATSCT-
SCAC, ca. 95% pure (CE), M 2602.0 (ESI-MS, calc. 2602.5),
were used for Tm measurements under conditions specified in
Table 2.
10 V. Tereshko, G. Minasov and M. Egli, J. Am. Chem. Soc., 1999, 121,
470 (Nucleic Acid Database BD0007).
11 F. Mohamadi, N. G. J. Richards, W. C. Guida, R. Liskamp,
M. Lipton, C. Caulfield, G. Chang, T. Hendrickson and W. C. Still,
J. Comput. Chem., 1990, 19, 440 www.schroedinger.com.
12 Gaussian 98 (Revision A.7), M. J. Frisch, G. W. Trucks,
H. B. Schlegel, G. E. Scuseria, M. A. Robb, J. R. Cheeseman, V. G.
Zakrzewski, J. A. Montgomery Jr., R. E. Stratmann, J. C. Burant,
S. Dapprich, J. M. Millam, A. D. Daniels, K. N. Kudin,
M. C. Strain, O. Farkas, J. Tomasi, V. Barone, M. Cossi, R. Cammi,
B. Mennucci, C. Pomelli, C. Adamo, S. Clifford, J. Ochterski,
G. A. Petersson, P. Y. Ayala, Q. Cui, K. Morokuma, D. K. Malick,
A. D. Rabuck, K. Raghavachari, J. B. Foresman, J. Cioslowski,
J. V. Ortiz, A. G. Baboul, B. B. Stefanov, G. Liu, A. Liashenko,
P. Piskorz, I. Komaromi, R. Gomperts, R. L. Martin, D. J. Fox,
T. Keith, M. A. Al-Laham, C. Y. Peng, A. Nanayakkara,
C. Gonzalez, M. Challacombe, P. M. W. Gill, B. G. Johnson,
W. Chen, M. W. Wong, J. L. Andres, M. Head-Gordon,
E. S. Replogle and J. A. Pople, Gaussian, Inc., Pittsburgh PA, 1998;
www.gaussian.com.
Acknowledgements
Per-Ola Norrby, The Technical University of Denmark, is
thanked for help with modeling, Britta M. Dahl for oligo-
nucleotide synthesis, Jette Poulsen and Anette W. Jørgensen
for technical assistance, and Cureon A/S for ESI-MS
measurements.
13 X. Chen, R. Guo and Z. Zhou, Acta Crystallogr. Sect. E, 2002, 58,
References
568.
1 C. Simons, Nucleoside Mimetics: Their Chemistry and Biological
Properties, Gordon and Breach Science Publishers, Amsterdam,
2001.
14 M. R. Harnden, P. G. Wyatt, M. R. Boyd and D. Sutton, J. Med.
Chem., 1990, 33, 187.
15 J. Zhou and P. B. Shevlin, Synth. Commun., 1997, 3591.
O r g . B i o m o l . C h e m . , 2 0 0 3 , 1, 3 2 9 3 – 3 2 9 6
3296