Soft metal-mediated base pairing with novel synthetic nucleosides possessing an O,S-donor ligand

Article abstract of DOI:10.1021/jo800587d

(Chemical Equation Presented) Metal-mediated base pairing with artificial ligand-bearing nucleosides allows site-selective metal incorporation inside DNA duplexes. In particular, this strategy has provided a general way of discrete, heterogeneous metal arrays in a programmable manner. To increase the kind of metallo-building blocks, we have newly synthesized two artificial nucleosides which have an O,S-donor ligand as the nucleobase moiety, mercaptopyridone (M) and hydroxypyridinethione (S). These nucleosides were found to efficiently form metal-mediated base pairs with soft transition metal ions such as Pd ²⁺ and Pt²⁺.

Full text of DOI:10.1021/jo800587d

Soft Metal-Mediated Base Pairing with Novel Synthetic Nucleosides
Possessing an O,S-Donor Ligand
Yusuke Takezawa,† Kentaro Tanaka,‡ Maiko Yori,† Shohei Tashiro,† Motoo Shiro,§ and
Mitsuhiko Shionoya*,†
Department of Chemistry, Graduate School of Science, The UniVersity of Tokyo, Hongo, Bunkyo-ku, Tokyo
113-0033, Japan, Department of Chemistry, Graduate School of Science, Nagoya UniVersity, Furo-cho,
Chikusa-ku, Nagoya 464-8602, Japan, and Rigaku Corporation, 3-9-12 Matsubara-cho, Akishima,
Tokyo 196-8666, Japan
shionoya@chem.s.u-tokyo.ac.jp
ReceiVed March 14, 2008
Metal-mediated base pairing with artificial ligand-bearing nucleosides allows site-selective metal
incorporation inside DNA duplexes. In particular, this strategy has provided a general way of discrete,
heterogeneous metal arrays in a programmable manner. To increase the kind of metallo-building blocks,
we have newly synthesized two artificial nucleosides which have an O,S-donor ligand as the nucleobase
moiety, mercaptopyridone (M) and hydroxypyridinethione (S). These nucleosides were found to efficiently
form metal-mediated base pairs with soft transition metal ions such as Pd2+ and Pt2+.
Introduction
acidity, metallo-base pairs have been expected to provide novel
structures and functions for artificial DNAs.
Incorporation of artificial base pairs into DNA double helices
has shown promise not only for expansion of genetic informa-
tion1 but also for construction of functional molecules.2 So far,
a number of excellent examples have been reported on artificial
base pairs through alternative hydrogen-bonding,3 hydrophobic
packing interactions,4 and metal-coordination.5–18 In particular,
because metal complexes have unique chemical and physical
properties such as redox-, optical-, magnetic-functions and Lewis
Since we reported Pd2+-mediated base pairing6 between two
o-phenylenediamine-bearing nucleosides, we6–9 and others10–15
have demonstrated artificial ligand-type nucleosides that form
(4) For example: (a) Schweitzer, B. A.; Kool, E. T. J. Am. Chem. Soc. 1995,
117, 1863–1872. (b) Leconte, A. M.; Chen, L.; Romesberg, F. E. J. Am. Chem.
Soc. 2005, 127, 12470–12471. (c) Hirao, I.; Kimoto, M.; Mitsui, T.; Fujiwara,
T.; Kawai, R.; Sato, A.; Harada, Y.; Yokoyama, S. Nat. Methods 2006, 3, 729–
735. (d) Hirao, I.; Mitsui, T.; Kimoto, M.; Yokoyama, S. J. Am. Chem. Soc.
2007, 129, 15549–15555.
(5) (a) Shionoya, M.; Tanaka, K. Curr. Opin. Chem. Biol. 2004, 8, 592–
597. (b) Tanaka, K.; Shionoya, M. Chem. Lett. 2006, 35, 695–699. (c) Tanaka,
K.; Shionoya, M. Coord. Chem. ReV. 2007, 251, 2732–2742. (d) Clever, G. H.;
Kaul, C.; Carell, T. Angew. Chem., Int. Ed. 2007, 46, 6226–6236.
(6) Tanaka, K.; Shionoya, M. J. Org. Chem. 1999, 64, 5002–5003.
(7) (a) Cao, H.; Tanaka, K.; Shionoya, M. Chem. Pharm. Bull. 2000, 48,
1745–1748. (b) Tanaka, K.; Tasaka, M.; Cao, H.; Shionoya, M. Eur. J. Pharm.
Sci 2001, 13, 77–83. (c) Tasaka, M.; Tanaka, K.; Shiro, M.; Shionoya, M.
Supramol. Chem. 2001, 13, 671–675. (d) Tanaka, K.; Tasaka, M.; Cao, H.;
Shionoya, M. Supramol. Chem. 2002, 14, 255–261. (e) Shionoya, M.; Tanaka,
K. Bull. Chem. Soc. Jpn. 2000, 73, 1945–1954.
* To whom correspondence should be addressed. Phone and fax: +81-3-
5841-8061.
† The University of Tokyo.
‡ Nagoya University.
§ Rigaku Corporation.
(1) (a) Henry, A. A.; Romesberg, F. E. Curr. Opin. Chem. Biol. 2003, 7,
727–733. (b) Hirao, I. Curr. Opin. Chem. Biol. 2006, 10, 622–627, and references
therein.
(2) For example: (a) Bergstorm, D. E. In Current Protocols in Nucleic Acid
Chemistry; Beaucage, S. L., Bergstorm, D. E., Glick, G. D., Jones, R. A. Eds.;
Wiley: New York, 2001; Unit 1.4. (b) Kool, E. T. Acc. Chem. Res. 2002, 35,
936–943. (c) Okamoto, A. Bull. Chem. Soc. Jpn. 2005, 78, 2083–2097.
(3) For example: (a) Switzer, C.; Moroney, S. E.; Benner, S. A. J. Am. Chem.
Soc. 1989, 111, 8322–8323. (b) Piccirilli, J. A.; Krauch, T.; Moroney, S. E.;
Benner, S. A. Nature 1990, 343, 33–37. (c) Sismour, A. M.; Benner, S. A. Nucleic
Acids Res. 2005, 33, 5640–5646.
(8) Tanaka, K.; Yamada, Y.; Shionoya, M. J. Am. Chem. Soc. 2002, 124,
8802–8803.
(9) Tanaka, K.; Tengeiji, A.; Kato, T.; Toyama, N.; Shiro, M.; Shionoya,
M. J. Am. Chem. Soc. 2002, 124, 12494–12498.
(10) Mu¨ller, J.; Bo¨hme, D.; Lax, P.; Cerda, M. M.; Roitzsch, M. Chem.-
Eur. J. 2005, 11, 6246–6253.
6092 J. Org. Chem. 2008, 73, 6092–6098
10.1021/jo800587d CCC: $40.75 2008 American Chemical Society
Published on Web 07/17/2008

NoVel Synthetic Nucleosides Possessing an O,S-Donor Ligand
CHART 1. Examples of Artificial Ligand-Type Nucleosides
CHART 2. Template-Directed Heterogeneous Metal Arrays
in Artificial DNA
synthesizer. Because oligonucleotides with any desired sequence
and length up to about 100 can be easily obtained by the
consecutive condensation of monomers, artificial nucleosides
can be similarly incorporated into DNA strands at the desired
positions in a designed sequence. In view of their possible
permutations, development of novel metallo-base pairs would
allow highly diverse metal arrays inside DNA, leading to the
functional fine-tuning of the metal complexes by predetermining
the sequence of ligand-type bases.
metal-mediated base pairs with Pd2+, Cu2+, Ag+, Ni2+, Mn3+
,
and so on. Some of them were successfully incorporated into
oligonucleotides in a programmable manner, and then provided
metal-mediated artificial base pairs mainly in duplexes. For
instance, we reported a pyridine-bearing nucleoside (P),8 which
forms a 2:1 linear complex with Ag+ or Hg2+, and a hydroxy-
pyridone-bearing nucleoside (H),9 which forms a 2:1 square-
planar complex with Cu2+ (Chart 1). Both nucleosides were
individually incorporated into DNA double strands and they
stabilized the duplexes through P-Ag+/Hg2+-P or H-
Cu2+-H base pairing. Moreover, the multiple incorporation of
nucleoside H allowed one-dimensional discrete assembly of
Cu2+ ions through H-Cu2+-H base pairing to form nCu2+ ·d(5′-
GHnC-3′)2 (n ) 1-5) inside the artificial DNA duplexes. The
Cu2+ ions stacked on top of each other were coupled ferro-
magnetically with one another through unpaired d electrons.16
Heterogeneous metal arrays of Cu2+ and Hg2+ were also
established by means of two nucleosides, P and H,17 in which
Toward this end it is essential to increase the kind of
nucleobase ligands that are specific to some metal ion(s). To
obtain well-defined metal-assembled complexes within DNA,
artificial metallo-base pairs should be designed so that their size
and shape go well with those of natural base pairs that are all
flat and stacked on top of each other. In light of the preferred
geometry of metallo-base pairs, linear-coordinating Ag+ and
Hg2+ and square-planar-coordinating Cu2+, low-spin Ni2+, Pd2+
,
and Pt2+ were first chosen from a series of transition metals. In
addition, the metal binding sites were placed at positions
corresponding to those of hydrogen bond donors or acceptors
of the natural base pairs. Both monodentate8,10 and bidentate
ligand-type nucleosides9,11 formed homogeneous metallo-base
pairs as two-coordinate linear and four-coordinate square-planar
complexes, respectively. Furthermore, heterogeneous base pair-
ing with [1 + 3]-coordination12,13 has been exploited using a
tridentate and a monodentate ligands. A salen-type nucleoside
also provided a base pair that is mediated by both metal
coordination and a reversible covalent cross-linking with
ethylenediamine.14
quantitative formation of Cu2+-Hg2+-Cu2+ and Cu2+-Cu2+
-
Hg2+-Cu2+-Cu2+ were achieved using predesigned artificial
DNA strands, d(5′-GHPHC-3′) and d(5′-GHHPHHC-3′), re-
spectively (Chart 2).
The selectivity in metal complexation is closely related to
the property of donor atoms of ligands as well as the kind of
metals. In this regard, the “hard and soft acids and bases”
(HSAB) rule, that hard (soft) ligands tend to bind to hard (soft)
metal ions, is a useful standpoint when we chose a proper
combination of donor atoms and metals.19 Artificial nucleosides
developed in this study were designed based on this rule, and
directed toward the site-selective, heterogeneous incorporation
of metals into DNA. Herein we describe the syntheses of novel
nucleosides, mercaptopyridone-bearing nucleoside (M) and
hydroxypyridinethione-bearing nucleoside (S), possessing O,S-
donor atoms as a bidentate ligand. These novel nucleosides were
found to form novel base pairs with soft metal ions such as
Pd2+ and Pt2+. Comparison of these nucleosides with hydroxy-
pyridone-bearing nucleoside (H) is also described.
The principal advantage of DNA synthesis is its “bottom-
up” protocol and straightforward preparation by automated DNA
(11) (a) Weizman, H.; Tor, Y. J. Am. Chem. Soc. 2001, 123, 3375–3376.
(b) Zhang, L.; Meggers, E. J. Am. Chem. Soc. 2005, 127, 74–75. (c) Switzer,
C.; Sinha, S.; Kim, P. H.; Heuberger, B. D. Angew. Chem., Int. Ed. 2005, 44,
1529–1532. (d) Switzer, C.; Shin, D. Chem. Commun. 2005, 1342–1344.
(12) (a) Meggers, E.; Holland, P. L.; Tolman, W. B.; Romesberg, F. E.;
Schultz, P. G. J. Am. Chem. Soc. 2000, 122, 10714–10715. (b) Atwell, S.;
Meggers, E.; Spraggon, G.; Schultz, P. G. J. Am. Chem. Soc. 2001, 123, 12364–
12367. (c) Zimmermann, N.; Meggers, E.; Schultz, P. G. Bioorg. Chem. 2004,
32, 13–25. (d) Shin, D.; Switzer, C. Chem. Commun. 2007, 4401–4403. (e)
Heuberger, B. D.; Shin, D.; Switzer, C. Org. Lett. 2008, 10, 1091–1094.
(13) Zimmermann, N.; Meggers, E.; Schultz, P. G. J. Am. Chem. Soc. 2002,
124, 13684–13685.
(14) (a) Clever, G. H.; Polborn, K.; Carell, T. Angew. Chem., Int. Ed. 2005,
44, 7204–7208. (b) Clever, G. H.; So¨ltl, Y.; Burks, H.; Spahl, W.; Carell, T.
Chem.-Eur. J. 2006, 12, 8708–8718. (c) Clever, G. H.; Carell, T. Angew. Chem.,
Int. Ed. 2007, 46, 250–253.
(15) (a) Brotschi, C.; Leumann, C. J. Nucleosides Nucleotides Nucleic Acids
2003, 22, 1195–1197. (b) Polonius, F.-A.; Mu¨ller, J. Angew. Chem., Int. Ed.
2007, 46, 5602–5604.
Results and Discussion
(16) Tanaka, K.; Tengeiji, A.; Kato, T.; Toyama, N.; Shionoya, M. Science
2003, 299, 1212–1213.
Molecular Design of Novel Nucleosides. We designed
mercaptopyridone-bearing nucleoside (M) and hydroxypyridi-
nethione-bearing nucleoside (S) having a thiol group and a
thiocarbonyl group, respectively, as a soft donor atom (Chart
1). These nucleosides were expected to form [2 + 2]-type base
(17) (a) Tanaka, K.; Clever, G. H.; Takezawa, Y.; Yamada, Y.; Kaul, C.;
Shionoya, M.; Carell, T. Nat. Nanotech. 2006, 1, 190–194. See also: (b) Mu¨ller,
J. Nature 2006, 444, 698.
(18) It is well-known that two thymine bases in the deprotonated form
coordinate to an Hg2+ ion to form T-Hg2+-T metallo-base pair. See: (a)
Kukleynik, Z.; Marzilli, L. G. Inorg. Chem. 1996, 35, 5654–5662. (b) Miyake,
Y.; Togashi, H.; Tashiro, M.; Yamaguchi, H.; Oda, S.; Kudo, M.; Tanaka, Y.;
Kondo, Y.; Sawa, R.; Fujimoto, T.; Machinami, T.; Ono, A. J. Am. Chem. Soc.
2006, 128, 2172–2173.
(19) Okada, T.; Tanaka, K.; Shiro, M.; Shionoya, M. Chem. Commun. 2005,
1484–1486.
J. Org. Chem. Vol. 73, No. 16, 2008 6093

Takezawa et al.
SCHEME 1. Synthetic Route for Mercaptopyridone-Bearing Nucleoside (M)a
a Reagents and conditions: (a) (i) PhLi, THF, -78 °C f 0 °C; (ii) S8, THF, -40 °C f -20 °C; (iii) Ph2NCOCl, THF, -78 °C f 0 °C, 55%; (b) TMSI,
CH3CN, reflux, quant.; (c) 1′-R-chloro-3′,5′-di-O-toluoyl-2′-deoxyribose, iPr2EtN, CH2Cl2, rt, 23%; (d) K2CO3, CH2Cl2/CH3OH, rt, 87%; (e) saturated aqueous
NH4OH, DTT, 90 °C, 31%.
pairs with softer transition metals in contrast to Cu2+-mediated
base pairing with hydroxypyridone-bearing nucleoside (H).9
Several examples of metal complexation of O,S-donor ligands
have been reported so far, such as N-substituted 3-hydroxy-2-
methyl-4-pyridinethione20,21 and 3-hydroxy-2-methyl-4-thiopy-
rone (thiomaltol).21,22 Lewis et al. reported that thiomaltol forms
a 2:1 complex with Ni2+ having a cis square-planar geometry,
and thereby concluded that the ligand exhibits a strong trans
FIGURE 1. Illustrations of nuclear Overhauser enhancements for two
influence.22 This tendency indicates that nucleosides possessing
anomeric isomers of 3.
O,S-donor ligands are likely to form square-planar metallo-base
pairs with a cis geometry that could fit well to DNA duplexes.
In addition, when these nucleosides, M and S, form a 2:1 base
pair with a divalent metal ion, metallo-base pairs become neutral
due to spontaneous deprotonation of the ligand upon metal
coordination. Therefore, these artificial base pairs were expected
to be rather stable due to hydrophobic effects within DNA
helices.
anomeric mixtures were separated by silica gel column chro-
matography to afford the desired ꢀ-nucleoside in 23%. Although
there was a possibility that not only desired N-nucleosides but
also O-nucleosides were obtained in the coupling reaction, the
formation of N-nucleoside was confirmed by its 13C NMR
spectrum that showed a signal of a carbonyl carbon at the 4
position of pyridone ring. The anomeric configuration of 3 was
determined by proton nuclear Overhauser effects (NOE) (Figure
1). When the 2′R proton resonances around 2.72 ppm were
Synthesis of Mercaptopyridone-Bearing Nucleoside (M).
Mercaptopyridone-bearing nucleoside (M) was synthesized
according to Scheme 1. The introduction of a protected thiol
separately irradiated, a nuclear Overhauser enhancement of 8%
functionality into the nucleobase framework was carried out by
was observed at the 1′ proton. The irradiation of the 2′ꢀ proton
gave a 10% enhancement at the 3′ proton, whereas only 1%
enhancement was observed at the 1′ proton. Furthermore, the
irradiation of the 1′ proton gave 5% enhancement at the 2′R
proton and 5% at the 4′ proton. These NOE trends provide a
clear evidence for ꢀ-anomer 3-ꢀ. The NOE measurement of
R-anomer 3-r was also carried out and gave reasonable results.
Deprotection of the toluoyl groups at the 3′ and 5′ positions
of nucleoside 3 afforded nucleoside 4. To avoid spontaneous
generation of the corresponding disulfide, the deprotection of
the diphenylcarbamoyl group of 4 was performed in saturated
ammonia solution in the presence of 1,4-dithiothreitol (DTT)
at 90 °C for 24 h, and the desired thiol-functional nucleoside
M was successfully obtained.
regioselective lithiation23 at the C3 position of 4-methoxypy-
ridine24 followed by thiolation and protection with a diphenyl-
carbamoyl group. The structure of the resulting 1 was deter-
1
mined by X-ray diffraction analysis (Figure S1) as well as H
NMR spectroscopy. Subsequent demethylation with trimethyl-
silyl iodide afforded a 4-carbonyl compound 2. During the
demethylation, nitrogen bubbling was essential to prevent the
formation of an N-methyl substituted byproduct.
The coupling reaction between the protected nucleobase 2
and R-chloro-2-deoxyribose25 furnished the nucleside 3 as a
mixtrure of R- and ꢀ-anomers (R/ꢀ ) 9:4) in 73% yield. The
(20) Katoh, A.; Tsukahara, T.; Saito, R.; Ghosh, K. K.; Yoshikawa, Y.;
Kojima, Y.; Tamura, A.; Sakurai, H. Chem. Lett. 2002, 31, 114–115.
(21) (a) Lewis, J. A.; Cohen, S. M. Inorg. Chem. 2004, 43, 6534–6536. (b)
Monga, V.; Patrick, B. O.; Orvig, C. Inorg. Chem. 2005, 44, 2666–2677. (c)
Monga, V.; Thompson, K. H.; Yuen, V. G.; Sharma, V.; Patrick, B. O.; McNeill,
J. H.; Orvig, C. Inorg. Chem. 2005, 44, 2678–2688.
Synthesis of Hydroxypyridinethione-Bearing Nucleoside
(S). Hydroxypyridinethione-bearing nucleoside (S) was synthe-
sized from the precursor of hydroxypyridone-bearing nucleoside
(H)9 (Scheme 2). Conversion of the carbonyl group of hydroxy-
pyridone 5 to a thiocarbonyl group by phosphorus pentasulfide
(P4S10)26 afforded pyridinethione 6. The transformation of the
(22) (a) Lewis, J. A.; Puerta, D. T.; Cohen, S. M. Inorg. Chem. 2003, 42,
7455–7459. (b) Lewis, J. A.; Tran, B. L.; Puerta, D. T.; Rumberger, E. M.;
Hendrickson, D. N.; Cohen, S. M. Dalton Trans. 2005, 2588–2596.
(23) Tre´court, F.; Mallet, M.; Mengin, O.; Gervais, B.; Que´guiner, G.
Tetrahedron 1993, 49, 8373–8380.
(24) Lister, T.; Prager, R. H.; Tsaconas, M.; Wilkinson, K. L. Aust. J. Chem.
2003, 56, 913–916.
(26) Dash, B.; Dora, E. K.; Panda, C. S. Heterocycles 1982, 19, 2093–2098.
(27) In the IR spectrum of 6, no absorption peak was observed around 1620
cm-1 for the carbonyl group of hydroxypyridone (Figure S3, Supporting
Information).
(25) This compound was prepared according to an improved procedure in
which acetyl chloride was used as an HCl generator. See: (a) Rolland, V.; Kotera,
M.; Lhomme, J. Synth. Commun. 1997, 27, 3505–3511.
6094 J. Org. Chem. Vol. 73, No. 16, 2008

NoVel Synthetic Nucleosides Possessing an O,S-Donor Ligand
SCHEME 2. Synthetic Route for Hydroxypyridinethione-Bearing Nucleoside (S)a
a Reagents and conditions: (a) P4S10, iPr2EtN, CH3CN, rt, 75%; (b) K2CO3, CH3OH, rt, 94%.
FIGURE 2. (a) UV absorption changes of nucleoside M at various concentrations of K2PdCl4. (b) Plots of absorbance at 236 and 413 nm against
the ratio of Pd2+ to M. [M] ) 50 µM in 25 mM MOPS (pH ) 7.0) at 25 °C, l ) 1.0 cm.
functional group was confirmed by IR27 and electrospray
ionization-time-of-flight (ESI-TOF) mass spectra (found: m/z
) 364.07 (z ) 1), calculated for C15H19NO6SNa [M + Na]+:
364.08). The following deacetylation of 6 gave the desired
nucleoside S. The X-ray diffraction analysis of its single crystals
confirmed the regioselective introduction of the sulfur atom and
ensured that the anomeric configuration was conserved (Figure
S2).
Metal-Mediated Base Pairing of the Nucleosides. Metal-
mediated base pairing of mercaptopyridone-bearing nucleoside
M was examined by the photometric titration study in a 3-(N-
morpholino)propanesulfonate (MOPS) buffer (pH 7.0) at 25 °C.
Complexation of M with Pd2+ took about 30 min, and with an
increase in Pd2+ concentrations, a new absorption peak at 413
nm appeared and the spectra changed linearly until the ratio of
[Pd2+]/[M] reached 0.5 (Figure 2). An ESI-TOF mass spectrum
FIGURE 3. ESI-TOF mass spectrum of a 2:2:1 mixture of M,
NaHCO3, and K2PdCl4 in H2O-CH3CN (1:1) (bp: metallo-base pair,
of a 2:1 mixture of M and Pd2+ in H2O-CH3CN (1:1) showed
2M: corresponding disulfide).
a signal at m/z 589.80 which is in good agreement with the
theoretical data for [M2Pd]+ complexes (Figure 3; found:
589.80, calculated for [bp - e]+: 590.00). This result indicates
that a metal-mediated base pair, M-Pd2+-M, was stoichio-
metrically formed.
quantitatively formed. Its ESI-TOF mass spectrum also con-
firmed the Ni2+-mediated base pairing (Figure 6; found: 542.04,
calculated for [bp - e]+: 542.03).
The ability of hydroxypyridinethione-bearing nucleoside S
to form a metal-mediated base pair was then examined with
square-planar-coordinating metal ions. Complexation of S with
Pt2+ reached to its equilibrium within 24 h. With an increase in
Pt2+ concentrations, the absorption at 338 nm arising from the
pyridinethione moiety gradually decreased and a new peak at
413 nm spontaneously appeared with two isosbestic points
(Figure 7). The spectra continued to change linearly until the
ratio of [Pt2+]/[S] reached 0.5. This result indicates the sto-
ichiometric formation of the metallo-base pair, S-Pt2+-S. Its
ESI-TOF mass spectrum also provided evidence for the com-
plexation in the ratio of [Pt2+]/[S] ) 1:2 (Figure 8; found:
730.13, calculated for [bp - e]+: 730.08).
In the titration of the nucleoside M with Ni2+, complexation
was rapidly completed and a shoulder peak around 375 nm
appeared upon addition of Ni2+ (Figure 4). The spectra changed
1
linearly in the range of [Ni2+]/[M] from 0 to 0.3. In the H
NMR spectrum in the case of the [Ni2+]/[M] ratio of 0.33:1 in
D2O, the signals of the nucleobase moiety were extremely
broadened (Figure 5). These results indicate the quantitative
formation of the octahedral 3:1 Ni2+ complex, and therefore
the nucleoside M would provide a metal-mediated base triplet
motif in the triple-stranded DNA. Further addition of Ni2+
resulted in the appearance of new peaks around the aromatic
region. The spectra converged when the ratio of [Ni2+]/[M]
reached 1:2, which suggests that an M-Ni2+-M base pair with
a diamagnetic square-planar structure with low-spin Ni2+ was
The metal-mediated base pairing properties of other combina-
tion of the ligand-type nucleosides and metal ions were also
J. Org. Chem. Vol. 73, No. 16, 2008 6095

Takezawa et al.
FIGURE 4. (a) UV absorption changes of nucleoside M at various concentrations of NiCl2. (b) Plots of absorbance at 274, 305, and 375 nm
against the ratio of Ni2+ to M. [M] ) 50 µM in 25 mM MOPS (pH ) 7.0) at 25 °C, l ) 1.0 cm.
with M and S exhibited their absorption over 340 nm, whereas
the absorption peak of metallo-base pairs with H appeared
around 300 nm. Thus, UV-vis absorption measurement would
be an excellent way to identify site-selective metal complexation
in artificial metallo-DNA.
Conclusion
In this paper, we described the efficient syntheses of two
artificial nucleosides, M and S, possessing an O,S-donor ligand
that coordinate to soft transition metals. The photometric titration
experiments with metal ions showed that both nucleosides form
metal-mediated base pairs efficiently with square-planar rela-
FIGURE 5. 1H NMR spectra of (a) M, (b) M + 1.0 equiv of NaHCO3,
tively soft metals such as Pd2+, Ni2+, and Pt2+. Thus, both M
(c) (b) + 0.33 equiv of NiCl2, and (d) (b) + 0.50 equiv of NiCl2. [M]
and S have great potential to form soft metal-mediated base
pairs within DNA oligomers and to align d8 transition metal
ions in a controllable way. Furthermore, these nucleosides would
assemble a variety of metals heterogeneously in DNA duplexes
in combination with a Cu2+-selective H nucleoside in a
programmable fashion. Further investigation on the incorpora-
tion of the nucleosides into DNA strands is currently under way.
) 2 mM in D2O at 300 K.
Experimental Section
4-Methoxy-3-thiodiphenylcarbamoylpyridine (1). 4-Methoxy-
pyridine (2.35 g, 21.6 mmol) was dissolved in THF (100 mL) and
cooled to -78 °C. To the solution was added dropwise phenyl-
lithium (1.1 M in cyclohexane-diethyl ether, 40.0 mL). After stirring
at 0 °C for 2 h, the mixture was kept at -40 °C and sulfur powder
(1.30 g, 40.5 mmol) was then added. The suspended solution was
stirred vigorously at -20 °C for 20 h to prepare thiolate. The
reaction mixture was cooled again to -78 °C followed by the
addition of a precooled THF solution (50 mL) of diphenylcarbamoyl
chloride (10.2 g, 44.0 mmol) and gradually allowed to warm up to
room temperature. After stirring for 20 h, the reaction mixture was
quenched with H2O (200 mL) and extracted with CHCl3 (200 mL
× 3). The combined organic phase was dried over anhydrous
MgSO4 and then concentrated. The residue was purified by silica
gel column chromatography (CHCl3:CH3OH ) 50:1) to afford 1
FIGURE 6. ESI-TOF mass spectrum of a 2:2:1 mixture of M,
NaHCO3, and NiCl2 in H2O-CH3CN (1:1) (bp: metallo-base pair, 2M:
corresponding disulfide).
estimated qualitatively by titration experiments (see Supporting
Information). The results are summarized with the absorption
maxima of metallo-base pairs as shown in Table 1. While both
M and S formed base pairs efficiently with relatively soft metal
ions such as Pd2+, Ni2+, and Pt2+, hydroxypyridone-bearing
nucleoside (H) did not quantitatively form a base pair with such
softer ions under the same condition. This result suggests that
the incorporation of both H and the sulfur-containing nucleoside,
M or S, into artificial oligonucleotides may allow heterogeneous
metal arrays in DNA duplexes. Moreover, metallo-base pairs
1
(4.52 g, 13.4 mmol, 62.1%) as a pale brown oil. H NMR (500
MHz, CDCl3): δ 8.61 (br, 1H), 8.56 (s, 1H), 7.47-7.34 (m, 10H),
7.17 (br, 1H), 4.10 (s, 3H). 13C NMR (125 MHz, CDCl3): δ 166.0,
165.9, 156.8, 152.9, 141.3, 129.3, 127.5, 115.3, 106.9. 56.0. FT-
IR (ATR): 1675, 1574, 1491, 1481, 1277, 1145, 1029, 1015, 823,
815, 695, 615, 575 cm-1. HRMS (ESI) m/z: [M + Na]+ calcd for
C19H16N2O2SNa 359.0830, found 359.0833.
6096 J. Org. Chem. Vol. 73, No. 16, 2008

NoVel Synthetic Nucleosides Possessing an O,S-Donor Ligand
FIGURE 7. (a) UV absorption changes of nucleoside S at various concentrations of K2PtCl4. (b) Plots of absorbance at 338 and 413 nm against
the ratio of Pt2+ to S. [S] ) 250 µM in 25 mM MOPS (pH ) 7.0) at 25 °C, l ) 0.1 cm.
13C NMR (125 MHz, DMSO-d6): δ 175.4, 165.9, 143.8, 141.5,
137.5, 129.4, 129.1, 128.0, 127.6, 117.5, 116.7. FT-IR (ATR): 1672,
1627, 1489, 1271, 1141, 851, 756, 699, 613, 580, 533 cm-1. HRMS
(ESI) m/z: [M + Na]+ calcd for C18H14N2O2SNa 345.0674, found
345.0692.
Fully Protected Mercaptopyridone-Bearing Nucleoside 3. To
a solution of 2 (170 mg, 0.526 mmol) in CH2Cl2 (2.0 mL) were
added diisopropylethylamine (94 µL, 0.540 mmol) and 1′-R-chrolo-
3′,5′-di-O-toluoyl-2′-deoxyribose (243 mg, 0.626 mmol). As stirring
at room temperature, the suspended solution gradually turned to a
yellow clear solution. After 3.5 h, the reaction mixture was washed
with H2O (50 mL) and the aqueous layer was extracted with CHCl3
(50 mL × 2). The combined organic phase was dried over
anhydrous MgSO4 and then concentrated. Silica gel column
chromatography (CHCl3:CH3OH ) 1:0 - 50:1) of the residue
afforded ꢀ-anomer (3) (59.6 mg, 0.0735 mmol, 14%) as a colorless
FIGURE 8. ESI-TOF mass spectrum of a 2:2:1 mixture of S, NaHCO3,
foam, while R-anomer (144 mg) and the mixture of both anomers
and K2PtCl4 in H2O-CH3OH (1:1) (bp: metallo-base pair).
(65.9 mg) were also obtained. The mixture was purified by further
column chromatography to obtain ꢀ-anomer in 23% total yield. 1H
TABLE 1. Metal-Mediated Base Pairing Properties of Nucleosides
with Mercaptopyridone (M), Hydroxypyridinethione (S), and
Hydroxypyridone (H)
NMR (500 MHz, CDCl3): δ 7.99 (d, J ) 2.4 Hz, 1H), 7.92 (d, J
) 8.2 Hz, 2H), 7.90 (d, J ) 8.1 Hz, 2H), 7.53 (dd, J ) 2.4, 7.8
Hz, 1H), 7.36-7.25 (m, 12H), 7.23 (d, J ) 8.1 Hz, 2H), 6.39 (d,
J ) 7.8 Hz, 1H), 5.80 (dd, J ) 5.4, 8.7 Hz, 1H), 5.62 (d, J ) 6.1
Hz, 1H), 4.75 (dd, J ) 3.3, 12.3 Hz, 1H), 4.66 (dd, J ) 3.0, 12.3
Hz, 1H), 4.58 (dd, J ) 3.0, 4.8 Hz, 1H), 2.72 (ddd, J ) 1.3, 5.4,
14.3 Hz, 1H), 2.50 (ddd, J ) 6.1, 8.7, 14.3 Hz, 1H), 2.44 (s, 3H),
2.36 (s, 3H); 13C NMR (125 MHz, CDCl3): δ 176.8, 166.4, 166.1,
165.8, 144.8, 144.6, 142.3, 135.4, 130.1, 129.8, 129.5, 129.4, 129.2,
129.0, 127.3, 126.3, 126.1, 120.6, 118.4, 93.5, 83.5, 74.8, 64.1,
39.8, 21.8, 21.7. FT-IR (ATR): 1715, 1681, 1633, 1591, 1489, 1265,
1176, 1093, 830, 750, 694, 613 cm-1. HRMS (ESI) m/z: [M +
Na]+ calcd for C39H34N2O7SNa 697.1984, found 697.2009.
Protected Mercaptopyridone-Bearing Nucleoside 4. A solution
of 3 (567 mg, 0.825 mmol) and K2CO3 (252 mg, 1.82 mmol) in a
mixture of CH2Cl2 (5.0 mL) and CH3OH (5.0 mL) was stirred at
room temperature for 2 h. The reaction mixture was concentrated
and then the residue was purified by silica gel chromatography
(CHCl3:CH3OH ) 50:1 - 5:1) to obtain 4 (313 mg, 0.715 mmol,
base-pairing property with metal
ionsa (λmax of metallo-base pair)
nucleoside
Cu2+
Pd2+
Ni2+
Pt2+
M
sb c (325 nm) Quantitativelyc sd (375 nme) Partiallyc
,
(413 nm)
(340 nme)
S
sb f (400 nme) Quantitativelyf Quantitativelyf Quantitativelyf
,
(397 nm)
Partiallyf
(316 nm)
(400 nme)
Partiallyf
(308 nm)
(413 nm)
c g
,
H
Quantitatively
(303 nm)
Not formedf
a In 25 mM MOPS (pH ) 7.0) at 25 °C. b The formation of several
species was speculated because of their complicated spectral changes
with no isosbestic point. c [Nucleoside]
)
50 µM. d Quantitative
formation of the 2:1 complex could not be confirmed by photometric
titration experiments. e Shoulder peaks. f [Nucleoside]
g Reference 9.
) 250 µM.
3-Thiodiphenylcarbamoyl-4-pyridone (2). A solution of 1 (453
mg, 1.35 mmol) and trimethylsilyl iodide (210 µL, 1.48 mmol) in
acetonitrile (30 mL) was heated at reflux for 20 h. To remove methyl
iodide generated, N2 bubbling was continued during the reaction.
The reaction was quenched by addition of CH3OH (5 mL) and the
solvent was evaporated. Silica gel column chromatography (CHCl3:
CH3OH ) 100:1 - 10:1) of the residue afforded 2 (444 mg, 1.38
1
87%) as a colorless foam. H NMR (500 MHz, CD3OD): δ 8.42
(d, J ) 2.3 Hz, 1H), 8.09 (dd, J ) 2.3, 7.6 Hz, 1H), 7.46-7.33
(m, 10H), 6.50 (d, J ) 7.6 Hz, 1H), 5.90 (dd, J ) 6.5, 6.5 Hz,
1H), 4.47-4.44 (m, 1H), 4.00 (dd, J ) 3.3, 6.4 Hz, 1H), 3.82 (dd,
J ) 3.2, 12.1 Hz, 1H), 3.75 (dd, J ) 3.2, 12.1 Hz, 1H), 2.45-2.40
(m, 1H), 2.38-2.33 (m, 1H). 13C NMR (125 MHz, CD3OD): δ
179.8, 168.3, 146.4, 139.8, 130.4, 129.3, 129.1, 120.0, 118.4, 95.4,
89.9, 72.0, 62.6, 43.2. FT-IR (ATR): 1678, 1630, 1561, 1489, 1268,
1
mmol) quantitatively as a pale yellow foam. H NMR (500 MHz,
DMSO-d6): δ 11.68 (br, 1H), 7.95 (s, 1H), 7.64 (d, J ) 7.2 Hz,
1H), 7.45-7.38 (m, 10H), 6.15 (d, J ) 7.1 Hz, 1H), 4.10 (s, 3H).
1180, 1145, 1095, 1056, 1002, 958, 830, 761, 698, 615 cm-1
.
J. Org. Chem. Vol. 73, No. 16, 2008 6097

Takezawa et al.
HRMS (ESI) m/z: [M + Na]+ calcd for C23H22N2O5SNa 461.1147,
found 461.1133.
Hydroxypyridinethione-Bearing Nucleoside (S). Compound 6
(0.16 g, 0.50 mmol) and K2CO3 (0.18 g, 1.3 mmol) were dissolved
in CH3OH (20 mL) and stirred at room temperature for 3 h. After
removal of the solvent, silica gel column chromatography (CHCl3:
CH3OH ) 10:1 - 1:1) afforded the desired nucleoside S (0.12 g,
0.46 mmol, 94%) as a yellow powder. 1H NMR (500 MHz,
CD3OD): δ 8.16 (d, J ) 7.0 Hz, 1H), 7.44 (d, J ) 7.0 Hz, 1H),
6.28 (dd, J ) 6.1, 6.1 Hz, 1H), 4.44 (m, 1H), 4.04 (m, 1H), 3.87
(dd, J ) 3.1, 12.2 Hz, 1H), 3.78 (dd, J ) 3.7, 12.2 Hz, 1H),
2.56-2.53 (m, 1H), 2.54 (s, 3H), 2.28 (m, 1H). 13C NMR (125
MHz, CD3OD): δ 171.1, 154.3, 128.5, 128.1, 126.8, 91.5, 89.9,
71.3, 62.2, 43.2, 12.3. MS m/z calcd for C11H15NO4S ([M + H]+):
258.08; found: 258.07. Anal. Calcd for C11H15NO4S: C, 51.35; H,
5.88; N, 5.44. Found: C, 51.12; H, 5.94; N, 5.24.
Mercaptopyridone-Bearing Nucleoside (M). Compound 4 (46.7
mg, 0.106 mmol) and 1,4-dithio-D,L-threitol (DTT, 59.6 mg, 0.386
mmol) were dissolved in saturated aqueous ammonia solution (8.0
mL) and heated in a screw-capped tube at 90 °C for 24 h. The
reaction mixture was concentrated and ammonia was removed in
vacuo. The residue was purified by reverse-phase silica gel column
chromatography (Wakogel 50C18, H2O). The fraction containing
the product was washed with AcOEt to remove DTT. The aqueous
layer was lyophilized to afford M (8.3 mg, 0.034 mmol, 32%) and
stored under nitrogen. Nucleoside M was stable under the condition
of the titration experiments without degassing the solvents, while
it was easily oxidized when heated. 1H NMR (500 MHz, CD3OD):
δ 8.37 (d, J ) 2.2 Hz, 1H), 8.07 (dd, J ) 2.2, 7.2 Hz, 1H), 6.58
(d, J ) 7.2 Hz, 1H), 5.94 (dd, J ) 6.5, 6.5 Hz, 1H), 4.45 (m, 1H),
4.03 (m, 1H), 3.83 (dd, J ) 3.2, 12.1 Hz, 1H), 3.76 (dd, J ) 3.5,
12.1 Hz, 1H), 2.45-2.43 (m, 1H), 2.37-2.31 (m, 1H). 13C NMR
(125 MHz, CD3OD): δ 175.0, 137.0, 136.0, 132.1, 111.9, 85.9,
90.1, 72.1, 62.7, 43.5. Anal. Calcd for C10H13NO4S·H2O: C, 45.97;
H, 5.79; N, 5.36. Found: C, 46.16; H, 5.66; N, 5.30.
Protected Hydroxypyridinethione-Bearing Nucleoside 6. Hy-
droxypyridone nucleoside 5 (2.87 g, 8.82 mmol) and P4S10 (2.01
g, 4.52 mmol) were dissolved in CH3CN (30 mL). To the
suspension was added dropwise a solution of diisopropylethylamine
(6.3 mL, 35 mmol) in CH3CN (40 mL) keeping it cold. The
resulting mixture was stirred for 4.5 h at room temperature followed
by addition of H2O (5 mL) with trapping generated H2S gas. After
evaporation of the solvent, the products were extracted with CHCl3
(100 mL × 3) from H2O (200 mL). The combined organic layer
was dried over MgSO4 and then concentrated in vacuo. Silica gel
column chromatography (CHCl3:CH3OH ) 100:1 - 10:1) afforded
the title compound (2.31 g, 6.77 mmol, 77%) as a brown-yellow
powder. 1H NMR (500 MHz, CDCl3): δ 8.73 (s, 1H), 7.63 (d, J )
7.1 Hz, 1H), 7.54 (d, J ) 7.1 Hz, 1H), 6.10 (dd, J ) 5.7, 7.7 Hz,
1H), 5.28 (ddd, J ) 2.4, 2.5, 6.5 Hz, 1H), 4.42 (m, 1H), 4.40 (m,
2H), 2.63 (ddd, J ) 2.5, 5.7, 14.3 Hz, 1H), 2.54 (s, 3H), 2.28 (m,
1H), 2.15 (s, 3H), 2.10 (s, 3H). 13C NMR (125 MHz, CDCl3): δ
173.3, 170.2, 170.1, 153.5, 126.1, 124.9, 124.7, 89.4, 83.4, 73.8,
63.3, 39.9, 20.9, 20.8, 12.5. FT-IR (ATR): 1735, 1601, 1447, 1429,
1382, 1363, 1337, 1234, 1199, 1107, 1032, 954, 878, 856, 653,
621 cm-1. HRMS (ESI) m/z: [M + Na]+ calcd for C15H19NO6SNa
364.0831, found: 364.0824.
Photometric Titration Experiments. The UV-vis spectra were
recorded at 25 °C. Solutions for titrations contained 50 µM of the
nucleoside M or 250 µM of S, and metal ions in 25 mM MOPS-
Na buffer (pH ) 7.0). Prior to the measurements, the samples
containing Pd2+ or Pt2+ ions were allowed to stand at room
temperature until the equilibrium was reached: approximately 30
min for Pd2+ and 24 h for Pt2+
.
ESI-TOF MS Measurements. The ESI-TOF mass spectra were
recorded on positive modes. The sample solutions were prepared
as follows: nucleoside (100 µM), K2PdCl4, NiCl2•6H2O or K2PtCl4
(50 µM), and NaHCO3 (100 µM) in H2O-CH3CN (1:1) or H2O-
CH3OH (1:1).
Acknowledgment. This work is supported by a Grant-in-
Aid for Exploratory Research to K.T. (No. 17655058), Scientific
Research (S) to M.S. (No. 16105001), and the Global COE
Program for Chemistry Innovation from the Ministry of Educa-
tion, Culture, Sports, Science and Technology (Japan), and
Grant-in-Aid for JSPS Fellows to Y.T. (No. 11921) from Japan
Society for the Promotion of Science (Japan).
Supporting Information Available: Characterization data
including 1H, 13C NMR spectra, NOE data, crystallographic data,
CIF files for 1 and S, UV absorption spectra of titration with
other combination of nucleosides and metal ions, and compara-
tive spectral data for competitive metal complexation. This
material is available free of charge via the Internet at
http://pubs.acs.org.
JO800587D
6098 J. Org. Chem. Vol. 73, No. 16, 2008