Pyrimidine-pyrimidine base pairs stabilized by silver(I) ions

Article abstract of DOI:10.1039/c0cc04091f

In the presence of Ag^I ions, the C-T and m⁵iC (5-methylisocytosine)-T base pairs showed comparable stability to the C-Ag ^I-C base pair, and the m⁵iC-C base pair was highly stabilized by the synergetic effect of

Full text of DOI:10.1039/c0cc04091f

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Pyrimidine–pyrimidine base pairs stabilized by silver(I) ionsw
Hidehito Urata,* Eriko Yamaguchi, Yasunari Nakamura and Shun-ichi Wada
Received 27th September 2010, Accepted 26th October 2010
DOI: 10.1039/c0cc04091f
In the presence of AgI ions, the C–T and m5iC (5-methyliso-
cytosine)–T base pairs showed comparable stability to the
C–AgI–C base pair, and the m5iC–C base pair was highly
stabilized by the synergetic effect of AgI coordination and
possible hydrogen bonding.
DNA forms
a double-stranded structure through the
formation of A–T and G–C Watson–Crick base pairs.1 The
selectivity of the hydrogen bonding between bases is essential
for the replication and expression of genetic information.
Recently, several types of artificial base pairing systems
formed by non-Watson–Crick type hydrogen bonding,2
hydrophobic interaction based on shape complementarity,3
and metal coordination4 have been reported. These non-
natural artificial bases form base pairs to stabilize duplex
structures. Among them, base pairs of the metal coordination
type can be used as a functional switching device that is
dependent on the presence or absence of metal ions. It has
been reported that T–T, U–U and C–C mismatch base pairs
are effectively stabilized by the addition of HgII and AgI ions,
respectively (Fig. 1).5–7 Ono and Togashi applied the metal-
ion-mediated base pairing system to detect HgII ions.8
Fig. 1 Structures of C–AgI–C, T–HgII–T and U–HgII–U base pairs.
In T–HgII–T and C–AgI–C base pairs, hydrogen bonding
cannot take place due to the non-complementarity of the T–T
and C–C base pairs. Recently, Muller and Polonius, however,
¨
reported a novel type of base pair involving both hydrogen
bonding and metal coordination through the Hoogsteen edge.9
Silver(I) ion has the ability to deprotonate thymine in oligo-
nucleotides and binds to the N3 atom of deprotonated thymine
to afford an AgI–thymine complex without any charge, and
this complex binds to the N7 position of 1-deazaadenine.9 This
suggests that the AgI–thymine complex has affinity toward an
aromatic tertiary nitrogen atom. Here, we describe the effects
of AgI ions on the stability of oligodeoxynucleotides (ODNs)
Fig. 2 Structures of pyrimidine–pyrimidine mismatch base pairs.
The ODNs used in this study were synthesized on an auto-
mated DNA synthesizer (Table 1).
Fig. 3 shows the UV melting curves of oligonucleotide
duplexes in the absence and presence of metal ions. In the
absence of metal ions (Fig. 3a), some of the curves are non-
sigmoidal and the Tm values were calculated by curve fitting
with the Meltwin program.12 Duplexes other than duplexes 2
and 4 showed similar Tm values (Table 1). The exceptional Tm
values for duplexes 2 and 4 may be attributed to their non-
sigmoidal transitions.
As reported by Ono and coworkers,5,7 the T–T and C–C
mismatch base pairs (duplexes 1 and 2) showed significant
stabilization when 1 equivalent of HgII and AgI ions,
respectively, was added (Fig. 3b, Table 1). It has been reported
that although AgI ion is able to deprotonate thymine to
form an AgI–thymine complex,9 it does not stabilize a T–T
mismatch base pair.5,13 To examine the binding affinity of the
AgI–thymine complex to cytosine, the effects of AgI ions on
the stability of the C–T mismatch base pair (duplex 3) were
evaluated. As shown in Fig. 3 and Table 1, the addition of AgI
containing
a pyrimidine–pyrimidine mismatch base pair
consisting of cytosine, thymine, and 5-methylisocytosine
(m5iC) (Fig. 2).
20-Deoxy-5-methylisocytidine, a more stable derivative of
20-deoxyisocytidine, was synthesized from thymidine according
to the literature procedure,10 with slight modifications.
20-Deoxy-5-methylisocytidine was protected with di(n-butyl)-
formamidine (dbf)11 for the 2-amino group and with
4,40-dimethoxytrityl group (DMT) for the 50-hydroxyl group,
and then converted into the phosphoramidite derivative.11
Osaka University of Pharmaceutical Sciences, 4-20-1 Nasahara,
Takatsuki, Osaka 569-1094, Japan. E-mail: urata@gly.oups.ac.jp;
Fax: +81 72-690-1089; Tel: +81 72-690-1089
w Electronic supplementary information (ESI) available: Synthetic
details of the duplexes containing m5iC and supplementary melting
curves. See DOI: 10.1039/c0cc04091f
c
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Chem. Commun., 2011, 47, 941–943 941

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Table 1 Synthesized duplexes containing a pyrimidine–pyrimidine
base pair mismatch and their melting temperatures (Tms)a
stabilized by the addition of AgI ions (DTm = 12.5 1C) and
the effects were slightly larger than those of duplex 3, although
duplex 3 was more stable than duplex 4 in the presence of AgI
ions. The m5iC–m5iC mismatch base pair is a structural
isoform of the C–C mismatch base pair and duplex 6 showed
thermal stability (36.2 1C) that was analogous to that of
duplex 2 (36.5 1C) in the presence of AgI ions. The relatively
large difference between the DTm values of duplexes 2 and 6
may be due to the non-sigmoidal transition in the melting
process of duplex 2 in the absence of AgI ions.
d(GA CGT X CTA CG)
(CT GCA Y GAT GC)d
Tm/1C
Tm/1C
(+ metal ion)
DTm/1C
X–Y
Duplex 1
Duplex 2
Duplex 3
Duplex 4
Duplex 5
Duplex 6
T–T
C–C
C–T
27.6
31.7
27.9
19.7
26.6
26.6
45.7b
36.5c
37.3c
32.2c
45.5c
36.2c
18.1
4.8
9.4
12.5
18.9
9.6
m5iC–T
m5iC–C
m5iC–m5iC
The m5iC–C base pair has two hydrogen bond donor–
acceptor pairs. In the presence of AgI ions, duplex 5 showed
a much higher Tm value (45.5 1C) than duplexes containing the
other AgI-mediated base pairs (32–37 1C). Notably, this Tm
value is comparable to that of the HgII-mediated T–T base
pair (45.7 1C) and is higher than that of the A–T full-match
sequence (X = A and Y = T, 45.0 1C) (not shown). This result
suggests the possibility that the AgI coordination and
hydrogen bonding synergistically contribute to the stabilization
of the m5iC–C mismatch base pair. As usual Ag–N bond length
is 2.1–2.2 A, the N3 to N3 distance of the m5iC–Ag–C complex
is approximately 4.2 A when the N3–Ag–N3 bond angle is 1801.
Thus, some deformation in the m5iC–C base pair such as buckle
or opening14 may be caused to form the hydrogen bond.
We have found that the AgI–thymine complex is able to
coordinate to cytosine and 5-methylisocytosine in ODNs.
These C–AgI–T and m5iC–AgI–T base pairs have roughly
comparable thermal stability to the C–AgI–C base pair. Thus,
the coordination of AgI ion would not be selective for the C–C
mismatch base pair. In addition, we have found that the
m5iC–AgI–C base pair is highly stable compared to the
C–AgI–C and C–AgI–T base pairs. This result demonstrated
the synergetic effect of metal coordination and possible
hydrogen bonding on the mismatch stabilization. These novel
silver(I)-mediated base pairs described here were stabilized
sufficiently by 1 equivalent of AgI ions (Fig. S1–S4, ESIw)
and thus the Kd values for the binding of AgI ions to these base
pairs would be less than 10 mM. Thus, AgI-coordinated
thymine and AgI-coordinated 5-methylisocytosine may be
useful as a recognition device for cytosine in ODNs. In
particular, the m5iC–AgI–C base pair may be a promising
device for sensing AgI ions.
a
Samples contained 5 mM duplex, 1 M NaClO4, and 10 mM MOPS,
pH 7.1. Tm values were calculated by using the Meltwin program.12
b
c
Tm value in the presence of 1 equivalent of HgII ions. Tm value in
the presence of 1 equivalent of AgI ions.
We thank Professor D. H. Turner and Dr J. A. McDowell
who generously provided the Meltwin program for calculation
of Tm values.
Fig. 3 UV melting curves of duplexes 1–6 in the absence (a) and
presence (b) of 1 equivalent of HgClO4 (duplex 1) or AgNO3
(duplexes 2–6). Samples contained 5 mM duplex, 1 M NaClO4, and
10 mM MOPS, pH 7.1.
Notes and references
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c
942 Chem. Commun., 2011, 47, 941–943
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c
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Chem. Commun., 2011, 47, 941–943 943