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CITATION: Toiviala, M., Kleemola, V., Maity, S., Lönnberg, T., Still 
Elusive – Pd(II)-Mediated Base Pairing by an 
Acetophenone Oxime Palladacycle within 15N-Labelled 
Double-Helical Oligonucleotides. European Journal of 
Organic Chemistry 2023, 26, e202201443. 
 
 which has been published in final form at 
 
DOI https://doi.org/10.1002/ejoc.202201443  
 
 
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1Still Elusive – Pd(II)-Mediated Base Pairing by an Acetophenone Oxime
Palladacycle within 15N-Labelled Double-Helical Oligonucleotides
Maaret Toivialaa, Vesa Kleemolaa, Sajal Maitya and Tuomas Lönnberga*
a Department of Chemistry, University of Turku, Henrikinkatu 2, 20500 Turku, Finland
Email: tuanlo@utu.fi
Abstract
α and β anomers of an acetophenone C-nucleoside were synthesized and incorporated in the middle
of short oligodeoxynucleotides. The ketone oligonucleotides were converted to 15N-labelled oxime
oligonucleotides by treatment with 15N-hydroxylamine and, finally, cyclopalladated by treatment with
lithium tetrachloropalladate. Comparison of the UV melting profiles of duplexes bearing the β anomer
of either the palladacyclic or the metal-free oxime C-nucleoside suggested formation of a stable Pd(II)-
mediated base pair, especially with adenine or thymine as the base pairing partner. Melting profiles
of the corresponding duplexes bearing the α anomer were much more convoluted, precluding
meaningful comparison. 15N NMR spectra were obtained for the β anomeric oxime oligonucleotide as
well as its palladacyclic derivative but the signals unfortunately diminished below detection limit when
the latter was hybridized with a complementary strand placing a 15N3-labelled thymine opposite to
the palladacyclic residue.
Keywords
oligonucleotides; palladium; base pairing; isotopic labeling; NMR spectroscopy
Introduction
Metal-mediated base pairing has been studied for more than two decades[1–8] for potential
applications ranging from molecular switches[9–11] and wires[12–16] to sequence recognition[17–21] to
2expansion of the genetic code[22–26]. Of the numerous metal ions used in these studies, Ag(I), Cu(II) and
Hg(II) have met with the greatest success. On the other hand, the metal ion employed in the very first
artificial metal-mediated base pair[27], namely Pd(II), has proven challenging and elusive to the
conventional experiments. Melting temperature measurements, for example, are complicated by the
relatively slow ligand exchange at Pd(II), which manifests itself as multiphasicity and hysteresis
between the denaturation and renaturation curves.[28–30] We have previously tried to address this
problem by complementing melting temperature measurements with a FRET-based strand
displacement assay, with moderate success.[30] However, strand displacement and melting
temperature experiments both fundamentally quantify the same property, namely hybridization
affinity of an oligonucleotide as a whole. Failure of the stability of a metal-mediated base pair to
translate into duplex stability of a corresponding oligonucleotide has been reported not only with
Pd(II)[31,32] but also with Ag(I)[33]. Such discrepancies are usually attributed to incompatible geometries
of the metal-mediated base pair and the double helix. An independent method for monitoring the
association and dissociation of a single metal-mediated base pair within a double-helical
oligonucleotide would, hence, be highly desirable.
15N NMR has already proven to be a valuable tool for studying Ag(I)-[34] and Hg(II)-[35,36]
mediated base pairing, although the insensitivity of 15N necessitates isotopic labeling of the ligands. In
this paper we report the first attempt at 15N NMR monitoring of Pd(II)-mediated base pairing within a
double-helical oligonucleotide. Acetophenone oxime palladacycle, allowing easy post-synthetic 15N-
labeling as well as comparison of the hybridization data with those obtained on related
oligonucleotides[29], was chosen as the Pd(II)-bearing nucleobase analogue. Both anomers of the
corresponding C-nucleoside were synthesized and studied separately.
Results and Discussion
Building block synthesis
Synthesis of the phosphoramidite building blocks of the α and β anomers of the acetophenone C-
nucleoside (1a and 1b, respectively) is outlined in Scheme 1. First, Wittig reaction between 2-deoxy-
3,5-O-[1,1,3,3-tetraisopropyl-1,3-disiloxanediyl]-D-erythro-pentose (2)[37] and 2-
(triphenylphosphoranylidene)acetophenone gave the enone intermediate 3 (a mixture of E and Z
isomers), as previously reported for related ketones and esters[38]. As a significant fraction of
compound 3 spontaneously cyclized to compound 4 (a mixture of α and β anomers), this reaction was
driven to completion by the addition of potassium tert-butoxide, rather than attempting to isolate
3compound 3. Removal of the Markiewicz protection by conventional fluoride treatment and
introduction of a 4,4´-dimethoxytrityl protection to the 5´-OH function afforded chromatographically
separable α and β anomers (6a and 6b, respectively) of the desired acetophenone C-nucleoside.
Absolute configuration of the anomeric carbon atom of 6a and 6b was established based on NOESY
couplings between the anomeric proton and either H3´ or H4´ (Figure 1). Finally, both anomers were
converted into phosphoramidite building blocks by conventional treatment with 2-cyanoethyl N,N-
diisopropylchlorophosphoramidite.
Scheme 1. Synthesis of the α and β anomers of an acetophenone C-nucleoside and the corresponding
protected phosphoramidite building blocks. Reagents and conditions: a) 2-
(triphenylphosphoranylidene)acetophenone, benzene, 50 C, 45 h; b) t-BuOK, THF, 25 C, 5 min; c)
Et3N3HF, THF, 25 C, 18 h; d) DMTrCl, pyridine, N2 atmosphere, 25 C, 16 h; e) 2-cyanoethyl N,N-
diisopropylchlorophosphoramidite, Et3N, CH2Cl2, N2 atmosphere, 25 C, 1 h.
OH
O
O
Si
Si
O
OH
O
O
Si
Si
O
O a
O
O
O
Si
Si
O
b
PhO
Ph
O
O
OH
HOc
PhO
O
OH
DMTrO
d
O
Ph
O
OH
DMTrO
O
Ph
O
O
DMTrO
O
Ph
O
O
DMTrO
O
Ph
P
O N
P
O N
NC
NC
1a
1b
2 3 4
e
e
5
6a
6b
4Figure 1. NOESY spectra of compounds 6a (A) and 6b (B), showing coupling of the anomeric proton
with either H3´ or H4´, respectively.
5Oligonucleotide synthesis
Sequences of the oligonucleotides used in this study are summarized in Table 1. The modified
oligonucleotides ON1ak and ON1bk, incorporating either the α or the β anomer of the ketone C-
nucleoside in the middle of the sequence, were synthesized on an automated DNA/RNA synthesizer
by the conventional phosphoramidite strategy. According to trityl response, coupling yields were
essentially quantitative throughout the sequence. On completion of chain assembly, the
oligonucleotides were deprotected and released from the solid support by the conventional
ammonolysis. The crude products were purified by RP-HPLC and characterized by ESI-TOF-MS.
Curiously, the desired products ON1ak and ON1bk eluted as two distinct peaks, possibly owing to
intramolecular imine formation between the ketone and an exocyclic amino function. In addition, two
faster-eluting peaks of comparable height were collected and found to contain oligonucleotides with
molecular weight 17 units higher than that of the desired products.
Table 1. Oligonucleotides used in this study.
Oligonucleotide Sequencea
ON1ak 5´-CGAGCaKCTGGC-3´
ON1bk 5´-CGAGCbKCTGGC-3´
ON1ao 5´-CGAGCaOCTGGC-3´
ON1bo 5´-CGAGCbOCTGGC-3´
ON1ao-Pd 5´-CGAGCaOPdCTGGC-3´
ON1bo-Pd 5´-CGAGCbOPdCTGGC-3´
ON2a 5´-GCCAGAGCTCG-3´
ON2c 5´-GCCAGCGCTCG-3´
ON2g 5´-GCCAGGGCTCG-3´
ON2t 5´-GCCAGTGCTCG-3´
ON2t* 5´-GCCAGT*GCTCG-3´
a aK and bK refer to the α and β anomers of the benzoylmethyl C-nucleoside, aO and bO to the
respective 15N-labeled oximes and aOPd and bOPd to their palladacycles. In each sequence, the residue
varied in the hybridization experiments has been underlined.
6The purified ketone oligonucleotides ON1ak and ON1bk were converted into the respective oxime
oligonucleotides ON1ao and ON1bo by treatment with aqueous 15N-hydroxylammonium chloride
(Scheme 2). On completion of the reaction, as verified by ESI-TOF-MS analysis, the product mixtures
were subjected to RP-HPLC purification. Two peaks with nearly identical retention times were
obtained, in all likelihood corresponding to E and Z isomers of the oxime. In contrast to ON1ak and
ON1bk, the abovementioned side products did not react with 15N-hydroxylammonium chloride,
suggesting a modification at the ketone function. A mass increase of 17 units would be consistent with
addition of ammonia, present in very high concentration during the final deprotection step.
Tentatively the corresponding imine could be unexpectedly stable during RP-HPLC purification but
ionize as the carbinolamine during MS analysis. It is, however, difficult to understand why the
ammonia would not be eventually displaced by hydroxylamine. No further attempts were made to
identify these side products.
Scheme 2. Oximation and cyclopalladation of the ketone oligonucleotides ON1ak and ON1bk.
Reagents and conditions: a) HO15NH3Cl, H2O, 25 C, 96 h; Li2PdCl4, NaOAc, H2O, 55 C, 96 h.
The slower-eluting isomers of the oxime oligonucleotides ON1ao and ON1bo were converted to the
respective palladacyclic oligonucleotides ON1ao-Pd and ON1bo-Pd by incubation in an aqueous
solution of lithium tetrachloropalladate and sodium acetate at 55 C for 4 d (Scheme 2). Configuration
of the oxime double bond of the starting materials was unknown but, according to previous reports[39],
both E and Z isomers eventually react to give the same palladacycle, with the oxime-OH and the
benzene ring trans to each other. RP-HPLC analysis of the product mixture revealed several new peaks
(chromatograms presented in the Supporting Information). This phenomenon has been reported
before[28–30,40,41] and attributed to relatively slow ligand exchange at Pd(II). The collected fractions were
analyzed by ESI-TOF-MS and those showing addition of a single Pd(II) ion combined. In line with
previous reports, the initially formed chlorido-bridged dimer could not be detected, suggesting
dissociation during HPLC purification.
7UV and CD melting studies
UV melting profiles of duplexes formed by the cyclopalladated oligonucleotides ON1ao-Pd and
ON1bo-Pd with the unmodified complementary strands ON2a, ON2c, ON2g and ON2t were measured
to assess the base pairing preferences of both anomers of the acetophenone oxime palladacycle C-
nucleoside within a double helix. Corresponding unmetallated duplexes were also studied for
reference. In the middle of each duplex, an artificial nucleobase surrogate (acetophenone oxime or
the respective palladacycle) was paired with a canonical nucleobase (adenine, cytosine, guanine or
thymine). The samples had an oligonucleotide concentration of 1.0 µM, pH of 7.4 (20 mM cacodylate
buffer) and ionic strength of 0.10 M (adjusted with sodium perchlorate).
Melting profiles of duplexes formed by the faster- and slower-eluting isomers of the
unmetallated oxime oligonucleotides ON1ao and ON1bo and the respective palladacyclic
oligonucleotides ON1ao-Pd and ON1bo-Pd with the unmodified complementary strand ON2a are
presented in Figure 2 and all melting profiles in the Supporting Informations (Figures S24—S47).
Melting curves of all duplexes incorporating the alpha anomer of the acetophenone oxime C-
nucleoside (Figure 2A) were multiphasic, making determination of melting temperatures somewhat
ambiguous and in some cases impossible. Furthermore, with the possible exception of ON1ao-
PdON2a, hybridization was not complete even at the low end of the experimental temperature range
(10 C). Figure 3 summarizes the highest melting temperatures associated with a significant transition
for all duplexes, while all melting temperatures are tabulated in Table 2. Overall, melting temperatures
of the “alpha duplexes” were highly scattered, precluding reasonable comparison or interpretation in
terms of probable binding modes.
In contrast to ON1ao, duplexes formed by both isomers of ON1bo showed typical monophasic
sigmoidal melting curves (Figure 2B), allowing reliable determination of a single melting temperature
for each duplex. These melting temperatures were remarkably similar irrespective of the isomer of
ON1bo or the canonical nucleobase opposite to the acetophenone oxime residue, ranging from 27 to
32 C (Figure 3). Melting curves of the respective palladacyclic duplexes, in turn, were biphasic
although the higher melting temperature could only be quantified with ON1bo-PdON2a and ON1bo-
PdON2t. Similar melting profiles have been previously reported for other duplexes incorporating
palladacyclic nucleobase surrogates[29,30] and the higher melting temperature proposed to stem from
dissociation of a Pd(II)-mediated base pair. A similar interpretation seems reasonable also in the
present case. With thymine as the base pairing partner, Pd(II) would in all likelihood coordinate to N3,
whereas with adenine both N1- and N7-coordination are possible.[42] At least duplexes ON1bo-
PdON2a and ON1bo-PdON2t would appear to be more stable than the previously studied[29]
8counterparts bearing either a more flexible or, especially, a more rigid analogue of the palladacyclic
oxime residue. Unfortunately, difficulties in determining the higher melting temperature in the latter
two cases preclude a more quantitative comparison.
Figure 2. UV melting profiles of duplexes A) ON1aoON2a and ON1ao-PdON2a and B) ON1boON2a
and ON1bo-PdON2a; pH = 7.4 (20 mM cacodylate buffer); [oligonucleotides] = 1.0 µM; I(NaClO4) =
0.10 M. In both panels, dotted and dashed lines refer to the faster- and slower-eluting isomers of the
unpalladated oxime oligonucleotide and solid line to the palladacyclic oligonucleotide.
20 40 60 80
1.00
1.05
1.10
1.15
1.20
1.25
A
 /
A
10
T / oC
20 40 60 80
1.00
1.05
1.10
1.15
A
 /
A
10
T / oC
A
B
9Figure 3. UV melting temperatures of duplexes formed by the modified oligonucleotides ON1ao,
ON1ao-Pd, ON1bo and ON1bo-Pd with the natural oligonucleotides ON2a (cyan), ON2c (magenta),
ON2g (yellow) and ON2t (black); pH = 7.4 (20 mM cacodylate buffer); [oligonucleotides] = 1.0 µM;
I(NaClO4) = 0.10 M. For each duplex, the column represents the highest melting temperature that
could be determined reliably. The error bars indicate standard errors of the average of three
experiments.
ON
1a
o (
fas
t)
ON
1a
o (
slo
w)
ON
1a
o-P
d
ON
1b
o (
fas
t)
ON
1b
o (
slo
w)
ON
1b
o-P
d
10
20
30
40
50
60
T m
 /
o C
10
Table 2. UV melting temperatures of the duplexes studied; pH = 7.4 (20 mM cacodylate buffer);
[oligonucleotides] = 1.0 µM; I(NaClO4) = 0.10 M. The error limits represent standard errors of the
average of three experiments.
ON2a ON2c ON2g ON2t
ON1ao (fast) 18.6  0.4 43.9  0.2 31.0  0.2
59.2  0.2
30.5  0.1
ON1ao (slow) 16.5  0.4
56.8  0.4
36.1  0.3 25.5  0.6
51.6  0.3
22.0  0.4
36.7  0.3
56.0  0.2
70.2  0.2
ON1ao-Pd 19.7  0.2
60.1  0.3
n.a.a 16.9  0.3
52  5
12.6  0.7
34.9  0.2
55.2  0.4
ON1bo (fast) 31.8  0.1 28.9  0.1 29.6  0.2 29.5  0.1
ON1bo (slow) 27.5  0.2 28.1  0.1 28.7  0.1 27.0  0.1
ON1bo-Pd 21.0  0.2
51.5  0.1
20.6  0.1 18.7  0.3 12  3
45  1
a No sigmoidal melting curve observed.
After completion of the UV melting studies, the samples were analyzed by UPLC-ESI-TOF-MS to verify
the stability of the palladacyclic oligonucleotides under the experimental conditions and to test
whether the putative duplexes could be detected directly. Signals for single-stranded ON1ao-Pd or
ON1bo-Pd and one of the complementary oligonucleotides were observed with each sample (Figure
S48—S55 in the Supporting Information), indicating that the palladacyclic oligonucleotides persisted
over several heating and cooling ramps. Duplexes, on the other hand, were not detected but this is
not very surprising given the denaturing conditions inside the UPLC column (T = 60 C and a significant
fraction of methanol in the eluent).
11
Secondary structures of the oligonucleotide duplexes were studied by CD spectropolarimetric
analysis of the same samples as used in the UV melting studies. The spectra were acquired at 10 C
intervals over a range of 10—90 C (Figure S56—S79 in the Supporting Information). With most
duplexes, the spectra obtained at 10 C resembled those of B-type double helices, with positive and
negative Cotton effects around 280 and 250 nm, respectively. With duplexes incorporating the α
anomer of the acetophenone oxime residue, the latter extended further towards shorter wavelengths
whereas with duplexes incorporating the β anomer, the two Cotton effects were more symmetric.
Duplexes ON1ao(faster-eluting isomer)ON2a, ON1ao(slower-eluting isomer)ON2c, ON1ao-
PdON2c and ON1bo(slower-eluting isomer)ON2t showed little ellipticity, suggesting a distorted
secondary structure. In all cases, both Cotton effects diminished on increasing temperature,
consistent with denaruration of the double helix.
For more quantitative comparison with the UV melting profiles, the molar ellipticity at 285 nm
was plotted as a function of temperature for each duplex (Figure S80—S103 in the Supporting
Information). In contrast to the UV melting profiles, the limited number of data points precluded
smoothing and Gaussian fitting of the first-derivative curve. Instead, Equation 1 was fitted to the
experimental data by a non-linear least-squares method, yielding a single melting temperature for
each duplex.
[𝜃]𝑜𝑏𝑠 = [𝜃]𝑠𝑠 + [𝜃]𝑑𝑠−[𝜃]𝑠𝑠1+10(𝑇𝑚−𝑇)𝑝 (1)
[]obs is the observed molar ellipticity, []ss and []ds molar ellipticities of the single- and double-
stranded oligonucleotides, Tm the melting temperature, T the measurement temperature and p the
slope of the sigmoidal curve. With the “alpha duplexes”, many of the CD melting temperatures were
highly uncertain but the more reliable ones were in reasonable agreement with the UV melting
temperatures corresponding to the most prominent transition. Curiously, CD melting temperatures of
most of the “beta duplexes” were consistently higher than the UV melting temperatures, by 15—30
C. A notable exception was duplex ON1bo-PdON2t, with which the two values were identical within
error limits.
15N NMR spectrometric studies
For 15N NMR spectrometric monitoring of the putative Pd(II)-mediated base pairing, ON1bo-Pd was
paired with ON2t*, placing a 15N3-labelled thymine opposite to the palladacyclic residue. 15N3-labelled
thymine was selected as the base pairing partner because of the synthetic availability of the
12
corresponding nucleoside phosphoramidite[43] and the unambiguous coordination preferences
(exclusively N3-coordination) of thymidine[42]. ON1bo-PdON2t was also one of the duplexes showing
a distinct high UV melting temperature (45  1 C), tentatively assigned to Pd(II)-mediated base
pairing. The measurements were carried out in D2O at pD of 7.8 (20 mM cacodylate buffer) and ionic
strength of 0.10 M (adjusted with NaClO4).
15N NMR spectrum of the slower-eluting isomer of the unpalladated oligonucleotide ON1bo
(Figure 4A) showed a peak at 349.2 ppm, characteristic of an aromatic oxime nitrogen[44]. Likewise,
the chemical shift of the sole peak in the spectrum of the complementary sequence ON2t* (Figure 4B,
156.0 ppm) was in good agreement with the previously reported values[35,36]. With the palladacyclic
oligonucleotide ON1bo-Pd, two peaks of nearly equal intensity were observed (Figure 4C, 351.0 and
360.7), possibly reflecting differences in the Pd(II) coordination sphere.
13
Figure 4. 15N NMR spectra of oligonucleotides A) ON1bo, B) ON2T* and C) ON1bo-Pd; T = 298 K; pD =
7.8 (20 mM cacodylate buffer in D2O); [oligonucleotides] = 150 µM; I(NaClO4) = 0.10 M.
Curiously, hybridization of equimolar amounts of ON1bo-Pd and ON2t* resulted in disappearance of
the 15N NMR signals of both oligonucleotides (data not shown). In the case of the complementary
14
strand ON2t*, this observation would be at least in qualitative agreement with signal broadening on
coordination of the paramagnetic Pd(II). Disappearance of the signals of ON1bo-Pd, on the other hand,
is more difficult to explain as in all likelihood the oxime nitrogen is coordinated to Pd(II) even before
hybridization. 15N—Pd(II)—15N coupling would also decrease the signal to noise ratio, possibly below
detection limit. 15N NMR spectrum of ON1bo-PdON2t* was recorded at 10 C intervals over a range
of 10—70 C to find out if the signals would reappear on dissociation of the duplex, but without
success (data not shown). It would be tempting to interpret these results as very high thermal stability
of the putative Pd(II)-mediated base pair but we feel that such a conclusion would be premature.
Conclusion
A short DNA oligonucleotide incorporating the β anomer of an acetophenone oxime C-nucleoside in
the middle of the sequence hybridized equally well with complementary oligonucleotides placing any
of the canonical nucleobases opposite to the modified residue, yielding a single well-defined UV
melting temperature. The respective palladacyclic duplexes, on the other hand, showed biphasic
melting curves, with one melting temperature lower and one higher than that of the unpalladated
duplex. Similar behavior has been reported previously for related duplexes and attributed to
sequential dissociation of the Watson—Crick and metal mediated parts. Duplexes bearing the α
anomer of the acetophenone oxime C-nucleoside, either in metal-free or palladacyclic form, gave
much more convoluted melting curves, in line with the “unnatural” orientation of the nucleobase
surrogate. The β anomer of the palladacyclic oligonucleotide gave two 15N NMR signals, presumably
corresponding to differences in the Pd(II) coordination sphere. Lamentably, these signals, as well as
that of the 15N3-labelled complementary sequence, diminished below detection limit on hybridization
of the two strands. While this result is consistent with a stable Pd(II)-mediated base pair, direct and
compelling NMR evidence still remains elusive.
Experimental
General methods
Unless otherwise stated, all chemicals, including the unmodified oligonucleotides, were commercial
products and used as received. Et3N was dried over CaH2 and solvents over activated 4 Å molecular
sieves. Freshly distilled Et3N was used for the preparation of HPLC elution buffers. NMR spectra were
recorded on a Bruker Biospin 500 MHz NMR spectrometer. The 1H and 13C chemical shifts (, ppm)
15
were referenced to the corresponding residual signal of the solvent and the 15N and 31P chemical shifts
to external liquid ammonia and phosphoric acid, respectively. Mass spectra were recorded on a Bruker
Daltonics micrOTOF-Q mass spectrometer.
2-[2-Deoxy-3,5-O-(1,1,3,3-tetraisopropyldisiloxan-1,3-diyl)-D-ribofuranosyl]acetophenone (4): 2-
Deoxy-3,5-O-(1,1,3,3-tetraisopropyldisiloxan-1,3-diyl)-D-ribofuranose (2, 1.250 g, 3.319 mmol) was
coevaporated from anhydrous benzene (2  5 mL). The residue and 2-
(triphenylphosphoranylidene)acetophenone (2.500 g, 6.572 mmol) were dissolved in anhydrous
benzene (100 mL) and the resulting mixture stirred at 50 C for 45 h. Hexane (20 mL) was added, the
white precipitate removed by filtration and the filtrate evaporated to dryness. The crude intermediate
3 thus obtained was dissolved in THF (8.9 mL) and KOtBu (50.0 mg, 0.446 mmol) was added. The
reaction mixture was stirred at 25 C for 5 min, after which saturated aqueous NH4Cl (100 mL) was
added. The resulting solution was extracted with a mixture EtOAc (30 mL) and hexane (70 mL), the
organic phase washed with brine (100 mL), dried over Na2SO4 and evaporated to dryness. The residue
was purified on a silica gel column eluting with a mixture of EtOAc and hexane (1:4, v/v), affording
0.9487 g (59%) of compound 4 (colorless glass) as a mixture of two anomers. 1H NMR (500 MHz, CDCl3,
major anomer) δ 7.98 (d, J = 7.8 Hz, 2H, Bz-H2 and H6), 7.59 (t, J = 7.4 Hz, 1H, Bz-H4), 7.48 (t, J = 7.6
Hz, 2H, Bz-H3 and H5), 4.66 (m, 1H, H1´), 4.45 (m, 1H, H3´), 4.05 (m, 1H, H5´), 3.81—3.72 (m, 2H, H4´
and H5´´), 3.45 (dd, J = 5.5, 21.3 Hz, 1H, BzCH2), 3.05 (dd, J = 7.1, 16.5 Hz, 1H, BzCH2), 2.29 (ddd, J =
4.9, 6.7, 12.7 Hz, 1H, H2´), 1.89 (m, 1H, H2´´), 1.09 (m, 28H, SiCH and SiCH(CH3)2). 13C NMR (126 MHz,
CDCl3, major anomer) δ 198.2 (Bz-CO), 136.9 (Bz-C1), 133.2 (Bz-C4), 128.6 (Bz-C3 and C5), 128.21 (Bz-
C2 and C6), 85.8 (C4´), 74.0 (C1´), 73.4 (C3´), 63.64 (C5´), 44.7 (BzCH2), 40.6 (C2´), 17.7—16.8
(SiCH(CH3)2), 13.6—12.5 (SiCH). 1H NMR (500 MHz, CDCl3, minor anomer) δ 7.98 (d, J = 7.8 Hz, 2H, Bz-
H2 and H6), 7.59 (t, J = 7.4 Hz, 1H, Bz-H4), 7.48 (t, J = 7.6 Hz, 2H, Bz-H3 and H5), 4.65 (m, 1H, H1´), 4.50
(q, J = 7.0 Hz, 1H, H3´), 4.01 (dd, J = 3.2, 11.4 Hz, 1H, H5´), 3.84 (m, 1H, H4´), 3.77 (m, 1H, H5´´), 3.48
(dd, J = 5.9, 20.9 Hz, 1H, BzCH2), 3.23 (dd, J = 7.4, 16.5 Hz, 1H, BzCH2), 2.56 (m, 1H, H2´), 1.89 (m, 1H,
H2´´), 1.09 (m, 28H, SiCH and SiCH(CH3)2). 13C NMR (126 MHz, CDCl3, major anomer) δ 197.8 (Bz-CO),
137.0 (Bz-C1), 133.2 (Bz-C4), 128.6 (Bz-C3 and C5), 128.18 (Bz-C2 and C6), 83.6 (C4´), 73.9 (C1´), 73.8
(C3´), 63.56 (C5´), 44.8 (BzCH2), 40.7 (C2´), 17.7—16.8 (SiCH(CH3)2), 13.6—12.5 (SiCH). HRMS (ESI+-
TOF): m/z calcd for [C25H42NaO5Si2]: 501.2463; found: 501.2451 [M + Na]+.
16
2-(2-Deoxy-D-ribofuranosyl)acetophenone (5): 2-[2-Deoxy-3,5-O-(1,1,3,3-tetraisopropyldisiloxan-
1,3-diyl)-D-ribofuranosyl]acetophenone (4, 0.9702 g, 2.026 mmol) was dissolved in THF (24.3 mL).
Et3N3HF (1.32 mL, 8.11 mmol) was added and the resulting mixture stirred at 25 C for 72 h. The
product mixture was purified on a silica gel column eluting with a mixture of MeOH and CH2Cl2 (8:92,
v/v), affording 0.3972 g (82%) of compound 5 (colorless glass) as a mixture of two anomers. 1H NMR
(500 MHz, CDCl3, major anomer) δ 7.95 (m, 2H, Bz-H2 and H6), 7.56 (m, 1H, Bz-H4), 7.45 (m, 2H, Bz-
H3 and H5), 4.69 (m, 1H, H1´), 4.35 (m, 1H, H3´), 3.89 (q, J = 4.0 Hz, 1H, H4´), 3.68 (m, 1H, H5´), 3.62
(dd, J = 4.9, 11.8 Hz, 1H, H5´´), 3.34 (dd, J = 6.4, 16.4 Hz, 1H, BzCH2), 3.19 (dd, J = 5.9, 16.6 Hz, 1H,
BzCH2), 2.18 (ddd, J = 2.4, 5.7, 13.1 Hz, 1H, H2´), 1.86 (ddd, J = 9.5, 6.5, 13.2 Hz, 1H, H2´´). 13C NMR
(126 MHz, CDCl3, major anomer) δ 198.5 (Bz-CO), 136.7 (Bz-C1), 133.44 (Bz-C4), 128.68 (Bz-C3 and C5),
128.3 (Bz-C2 and C6), 87.1 (C4´), 74.7 (C1´), 73.2 (C3´), 63.1 (C5´), 44.2 (BzCH2), 41.2 (C2´). 1H NMR (500
MHz, CDCl3, minor anomer) δ 7.95 (m, 2H, Bz-H2 and H6), 7.56 (m, 1H, Bz-H4), 7.45 (m, 2H, Bz-H3 and
H5), 4.69 (m, 1H, H1´), 4.36 (m, 1H, H3´), 3.95 (q, J = 4.6 Hz, 1H, H4´), 3.68 (m, 1H, H5´), 3.62 (dd, J =
4.9, 11.8 Hz, 1H, H5´´), 3.50 (dd, J = 7.0, 17.0 Hz, 1H, BzCH2), 3.22 (dd, J = 5.8, 17.5 Hz, 1H, BzCH2), 2.49
(m, 1H, H2´), 1.81 (m, 1H, H2´´). 13C NMR (126 MHz, CDCl3, minor anomer) δ 198.9 (Bz-CO), 136.8 (Bz-
C1), 133.40 (Bz-C4), 128.65 (Bz-C3 and C5), 128.2 (Bz-C2 and C6), 85.5 (C4´), 74.4 (C1´), 72.6 (C3´), 62.4
(C5´), 44.9 (BzCH2), 40.4 (C2´). HRMS (ESI+-TOF): m/z calcd for [C13H16NaO4]: 259.0941; found: 259.0943
[M + Na]+.
2-[2-Deoxy-5-O-(4,4´-dimethoxytrityl)-α-D-ribofuranosyl]acetophenone (6a) and 2-[2-Deoxy-5-O-
(4,4´-dimethoxytrityl)-β-D-ribofuranosyl]acetophenone (6b): 2-(2-Deoxy-D-
ribofuranosyl)acetophenone (5) was coevaporated from anhydrous pyridine (3  10 mL) and the
residue was dissolved in anhydrous pyridine (5 mL) under N2 atmosphere. A solution of DMTrCl
(0.6220 g, 1.849 mmol) in anhydrous CH2Cl2 (1 mL) was added and the resulting mixture stirred at 25
C under N2 atmosphere for 5 h, after which it was concentrated to approximately 33% volume. The
remaining solution was diluted with CH2Cl2 (100 mL), washed with saturated aqueous NaHCO3 (100
mL), dried over Na2SO4 and evaporated to dryness. The residue was purified on a silica gel column
eluting with a mixture of Et3N, MeOH and CH2Cl2 (1:4:95, v/v), affording 0.2319 g (25%) of the α
anomer 6a and 0.3614 g (39%) of the β anomer 6b (yellowish foam). 1H NMR (500 MHz, CD3CN, 6a) δ
8.04 (m, 2H, Bz-H2 and H6), 7.64 (m, 1H, Bz-H4), 7.54 (m, 2H, Bz-H3 and H5), 7.45 (m, 2H, Ph-H2 and
H6), 7.31 (m, 6H, MeOPh-H2 and H6 and Ph-H3 and H5), 7.24 (m, 1H, Ph-H4), 6.87 (m, 4H, MeOPh-H3
and H5), 4.63 (m, 1H, H1´), 4.24 (m, 1H, H3´), 3.93 (q, J = 4.5 Hz, 1H, H4´), 3.79 (s, 6H, ArOCH3), 3.50
(dd, J = 7.3, 16.0 Hz, 1H, BzCH2), 3.21 (dd, J = 5.7, 16.0 Hz, 1H, BzCH2), 3.17 (d, J = 4.6 Hz, 3´-OH), 3.06
17
(dd, J = 4.0, 10.0 Hz, 1H, H5´), 3.02 (dd, J = 5.4, 10.0 Hz, 1H, H5´´), 2.44 (m, 1H, H2´), 1.70 (ddd, J = 7.1,
5.6, 12.7 Hz, 1H, H2´´). 13C NMR (126 MHz, CD3CN, 6a) δ 198.8 (Bz-CO), 158.6 (MeOPh-C4), 145.3 (Ph-
C1), 136.17 (Bz-C1), 136.16 (MeOPh-C1), 133.1 (Bz-C4), 130.0 (MeOPh-C2 and C6), 128.7 (Bz-C3 and
C5), 128.2 (Bz-C2 and C6), 128.1 (Ph-C3 and C5), 127.8 (Ph-C2 and C6), 126.7 (Ph-C4), 113.0 (MeOPh-
C3 and C5), 85.8 (Ar3C), 84.7 (C4´), 75.1 (C1´), 73.0 (C3´), 64.5 (C5´), 54.9 (ArOCH3), 45.1 (BzCH2), 40.5
(C2´). 1H NMR (500 MHz, CD3CN, 6b) δ 7.98 (m, 2H, Bz-H2 and H6), 7.62 (m, 1H, Bz-H4), 7.50 (m, 2H,
Bz-H3 and H5), 7.46 (m, 2H, Ph-H2 and H6), 7.32 (m, 6H, MeOPh-H2 and H6 and Ph-H3 and H5), 7.24
(m, 1H, Ph-H4), 6.85 (m, 4H, MeOPh-H3 and H5), 4.63 (m, 1H, H1´), 4.22 (m, 1H, H3´), 3.81 (m, 1H,
H4´), 3.77 (s, 6H, ArOCH3), 3.41 (dd, J = 7.2, 16.3 Hz, 1H, BzCH2), 3.15 (dd, J = 5.6, 16.4 Hz, 1H, BzCH2),
3.13 (br, 1H, 3´-OH), 3.07 (dd, J = 4.3, 10.0 Hz, 1H, H5´), 3.01 (dd, J = 5.0, 9.9 Hz, 1H, H5´´), 2.06 (ddd, J
= 2.0, 5.5, 12.9 Hz, 1H, H2´), 1.84 (ddd, J = 6.2, 9.7, 13.1 Hz, 1H, H2´´). 13C NMR (126 MHz, CD3CN, 6b)
δ 198.4 (Bz-CO), 158.6 (MeOPh-C4), 145.3 (Ph-C1), 136.16 (Bz-C1), 136.15 (MeOPh-C1), 133.1 (Bz-C4),
130.0 (MeOPh-C2 and C6), 128.7 (Bz-C3 and C5), 128.1 (Bz-C2 and C6 and Ph-C3 and C5), 127.8 (Ph-
C2 and C6), 126.8 (Ph-C4), 113.0 (MeOPh-C3 and C5), 85.9 (C4´), 85.8 (Ar3C), 74.8 (C1´), 73.1 (C3´), 64.6
(C5´), 54.9 (ArOCH3), 44.5 (BzCH2), 40.8 (C2´). HRMS (ESI+-TOF): m/z calcd for [C34H34NaO6]: 561.2248;
found: 561.2259 [M + Na]+.
2-{2-Deoxy-5-O-(4,4´-dimethoxytrityl)-3-O-[(2-cyanoethoxy)-(N,N-diisopropylamino)phosphinyl]-α-
D-ribofuranosyl}acetophenone (1a): 2-[2-Deoxy-5-O-(4,4´-dimethoxytrityl)-α-D-
ribofuranosyl]acetophenone (6a, 0.2319 g, 0.4305 mmol) was dissolved in anhydrous CH2Cl2 (4.5 mL)
under N2 atmosphere. Et3N (300 µL, 2.15 mmol) and 2-cyanoethyl-N,N-
diisopropylchlorophosphoramidite (116 µL, 0.520 mmol) were added and the resulting mixture stirred
at 25 C under N2 atmosphere for 1 h. The product mixture was diluted with CH2Cl2 (20 mL), washed
with saturated aqueous NaHCO3 (20 mL), dried over Na2SO4 (20 mL) and evaporated to afford 0.2327
g (73%) of compound 1a (white foam) in sufficient purity for oligonucleotide synthesis. 1H NMR (500
MHz, CD3CN) δ 8.03 (m, 2H, Bz-H2 and H6), 7.64 (m, 1H, Bz-H4), 7.54 (m, 2H, Bz-H3 and H5), 7.45 (m,
2H, Ph-H2 and H6), 7.32 (m, 6H, MeOPh-H2 and H6 and Ph-H3 and H5), 7.24 (m, 1H, Ph-H4), 6.87 (m,
4H, MeOPh-H3 and H5), 4.72 (m, 1H, H1´), 4.48 (m, 1H, H3´), 4.11 (m, 1H, H4´), 3.79 (s, 3H, ArOCH3),
3.78 (s, 3H, ArOCH3), 3.77—3.63 (m, 2H, POCH2), 3.63—3.47 (m, 3H, PNCH and BzCH2), 3.23 (m, 1H,
BzCH2), 3.13 (m, 1H, H5´), 3.04 (m, 1H, H5´´), 2.61 (t, J = 6.0 Hz, 1H, NCCH2), 2.52 (m, 2H, NCCH2 and
H2´), 1.95—1-81 (m, 1H, H2´´), 1.17 (d, J = 6.5 Hz, 3H, NCH(CH3)2), 1.16 (d, J = 6.4 Hz, 3H, NCH(CH3)2),
1.13 (d, J = 6.8 Hz, 3H, NCH(CH3)2), 1.06 (d, J = 6.8 Hz, 3H, NCH(CH3)2). 13C NMR (126 MHz, CD3CN) δ
198.7 (Bz-CO), 158.6 (MeOPh-C4), 145.3 (Ph-C1), 137.39 (Bz-C1), 137.35 (Bz-C1), 136.10 (MeOPh-C1),
18
136.05 (MeOPh-C1), 136.04 (MeOPh-C1), 136.00 (MeOPh-C1), 133.1 (Bz-C4), 130.0 (MeOPh-C2 and
C6), 128.7 (Bz-C3 and C5), 128.14 (Bz-C2 and C6), 128.13 (Bz-C2 and C6), 128.07 (Ph-C3 and C5), 128.0
(Ph-C3 and C5), 127.8 (Ph-C2 and C6), 126.77 (Ph-C4), 126.75 (Ph-C4), 118.5 (CN), 118.4 (CN), 113.0
(MeOPh-C3 and C5), 85.9 (Ar3C), 84.2 (d, J = 3.8 Hz, C4´), 84.0 (d, J = 6.2 Hz, C4´), 75.54 (C1´), 75.47 (d,
J = 16.3 Hz, C3´), 75.1 (d, J = 17.1 Hz, C3´), 64.3 (C5´), 64.2 (C5´), 58.41 (d, J = 18.4 Hz, POCH2), 58.38 (d,
J = 19.0 Hz, POCH2), 54.92 (ArOCH3), 54.90 (ArOCH3), 44.87 (BzCH2), 44.85 (BzCH2), 42.9 (d, J = 12.4 Hz,
PNCH), 39.50 (d, J = 3.6 Hz, C2´), 39.47 (d, J = 4.6 Hz, C2´), 23.92 (NCH(CH3)2), 23.88 (NCH(CH3)2), 23.86
(NCH(CH3)2), 23.83 (NCH(CH3)2), 20.03 (d, J = 7.2 Hz, NCCH2), 19.97 (d, J = 7.1 Hz, NCCH2). 31P NMR (202
MHz, CD3CN) δ 147.6. HRMS (ESI+-TOF): m/z calcd for [C34H34NaO6]: 739.3507; found: 739.3541 [M +
H]+.
2-{2-Deoxy-5-O-(4,4´-dimethoxytrityl)-3-O-[(2-cyanoethoxy)-(N,N-diisopropylamino)phosphinyl]-α-
D-ribofuranosyl}acetophenone (1b): 2-[2-Deoxy-5-O-(4,4´-dimethoxytrityl)-α-D-
ribofuranosyl]acetophenone (6b, 0.1566 g, 0.2907 mmol) was dissolved in anhydrous CH2Cl2 (3.0 mL)
under N2 atmosphere. Et3N (202 µL, 1.45 mmol) and 2-cyanoethyl-N,N-
diisopropylchlorophosphoramidite (78 µL, 0.351 mmol) were added and the resulting mixture stirred
at 25 C under N2 atmosphere for 1 h. The product mixture was diluted with CH2Cl2 (20 mL), washed
with saturated aqueous NaHCO3 (20 mL), dried over Na2SO4 (20 mL) and evaporated to afford 0.1826
g (85%) of compound 1b (white foam) in sufficient purity for oligonucleotide synthesis. 1H NMR (500
MHz, CD3CN) δ 7.99 (m, 2H, Bz-H2 and H6), 7.63 (m, 1H, Bz-H4), 7.51 (m, 2H, Bz-H3 and H5), 7.46 (m,
2H, Ph-H2 and H6), 7.31 (m, 6H, MeOPh-H2 and H6 and Ph-H3 and H5), 7.23 (m, 1H, Ph-H4), 6.85 (m,
4H, MeOPh-H3 and H5), 4.64 (m, 1H, H1´), 4.45 (m, 1H, H3´), 3.98 (m, 1H, H4´), 3.80 (m, 1H, POCH2),
3.777 (s, 1.5H, ArOCH3), 3.774 (s, 1.5H, ArOCH3), 3.772 (s, 1.5H, ArOCH3), 3.769 (s, 1.5H, ArOCH3), 3.69
(m, 1H, POCH2), 3.59 (m, 2H, PNCH), 3.46 (m, 1H, BzCH2), 3.19 (dd, J = 5.3, 16.5 Hz, 1H, BzCH2), 3.14
(m, 1H, H5´), 3.02 (m, 1H, H5´´), 2.65 (t, J = 6.0 Hz, 1H, NCCH2), 2.54 (t, J = 6.0 Hz, 1H, NCCH2), 2.33—
2.19 (m, 1H, H2´), 1.92 (ddd, J = 6.2, 9.7, 13.1 Hz, 1H, H2´´), 1.19 (d, J = 6.9 Hz, 3H, NCH(CH3)2), 1.17 (d,
J = 5.7 Hz, 3H, NCH(CH3)2), 1.16 (d, J = 5.6 Hz, 3H, NCH(CH3)2), 1.07 (d, J = 6.8 Hz, 3H, NCH(CH3)2). 13C
NMR (126 MHz, CD3CN) δ 198.2 (Bz-CO), 158.6 (MeOPh-C4), 145.3 (Ph-C1), 137.3 (Bz-C1), 136.10
(MeOPh-C1), 136.04 (MeOPh-C1), 135.99 (MeOPh-C1), 133.1 (Bz-C4), 130.06 (MeOPh-C2 and C6),
130.04 (MeOPh-C2 and C6), 130.02 (MeOPh-C2 and C6), 128.7 (Bz-C3 and C5), 128.2 (Bz-C2 and C6),
128.10 (Bz-C2 and C6), 128.09 (Ph-C3 and C5), 128.04 (Ph-C3 and C5), 127.8 (Ph-C2 and C6), 126.77
(Ph-C4), 126.75 (Ph-C4), 118.6 (CN), 118.4 (CN), 113.0 (MeOPh-C3 and C5), 85.87 (Ar3C), 85.86 (Ar3C),
85.20 (d, J = 4.4 Hz, C4´), 84.95 (d, J = 5.5 Hz, C4´), 75.4 (d, J = 17.2 Hz, C3´), 75.2 (d, J = 16.9 Hz, C3´),
19
75.1 (C1´), 75.0 (C1´), 64.14 (C5´), 64.10 (C5´), 58.4 (d, J = 19.6 Hz, POCH2), 54.9 (ArOCH3), 44.27
(BzCH2), 44.25 (BzCH2), 43.0 (d, J = 12.3 Hz, PNCH), 39.9 (d, J = 3.6 Hz, C2´), 39.8 (d, J = 4.7 Hz, C2´),
23.93 (d, J = 3.6 Hz, NCH(CH3)2), 23.89 (d, J = 3.6 Hz, NCH(CH3)2), 23.87 (d, J = 3.6 Hz, NCH(CH3)2), 23.83
(d, J = 3.6 Hz, NCH(CH3)2), 20.06 (d, J = 7.3 Hz, NCCH2), 19.97 (d, J = 6.6 Hz, NCCH2). 31P NMR (202 MHz,
CD3CN) δ 147.3, 147.2. HRMS (ESI+-TOF): m/z calcd for [C34H34NaO6]: 739.3507; found: 739.3541 [M +
H]+.
Oligonucleotide synthesis
Oligonucleotides ON1ak and ON1bk were assembled in 40 µmol scale on polystyrene support by an
ÄKTA Oligopilot plus 10 DNA/RNA synthesizer. Conventional phosphoramidite strategy was followed,
with 5-(benzylthio)-1H-tetrazole as the activator. A recycling time of 180 s was employed for the
commercially available building blocks as well as the acetophenone C-nucleoside phosphoramidites
1a and 1b. The coupling yields, based on trityl response, were essentially quantitative throughout the
syntheses. After chain assembly, the oligonucleotide products were released from the solid supports
and the base and phosphate protections removed by incubation in 25 % aqueous NH3 at 55 C for 16
h. The crude products were purified by RP-HPLC on a Hypersil ODS C18 column (250  10 mm, 5 µm)
eluting with a linear gradient (10—40% over 25 min) of MeCN in aqueous triethylammonium acetate,
the flow rate being 3.0 mL min-1 and the detection wavelength 260 or 290 nm (for the largest
injections, a wavelength offset from the absorption maximum of the oligonucleotides was used to
avoid saturation of the detector). HPLC traces are presented in Figures S15A and S16A of the
Supporting Information. The identity of the purified oligonucleotides was established by ESI-TOF-MS
and the quantity by UV spectrophotometry. The UV spectra were recorded over a range of 200—400
nm with a Jenway Genova Nano micro-volume spectrophotometer and the molar absortivities of the
oligonucleotides were calculated by an implementation of the nearest-neighbors method, assuming
the absorptivity of the acetophenone residue to be negligible.
The ketone oligonucleotides ON1ak and ON1bk were converted to the corresponding 15N-
labelled oxime oligonucleotides ON1ao and ON1bo by incubation in 50 mM aqueous 15N-
hydroxylammonium chloride at 25 C for 96 h. The product mixtures were fractioned by RP-HPLC
(Figures S15B and S16B of the Supporting Information) and the purified products characterized and
quantified as described above. Two peaks of nearly equal retention times and identical mass spectra
were obtained for both ON1ao and ON1bo, most likely corresponding to E and Z isomers of the oxime
moiety.
20
Cyclopalladation of the oxime oligonucleotides ON1ao and ON1bo was carried out by
incubation in an aqueous solution of Li2PdCl4 and NaOAc at 55 C for 96 h. Concentrations of the
oligonucleotide, Li2PdCl4 and NaOAc were 4.0 mM, 6.0 mM and 24 mM, respectively. The product
mixtures were fractioned by RP-HPLC (Figure S17 of the Supporting Information) and the purified
products characterized and quantified as described above. Several peaks showing the expected mass
spectrum were observed, in line with previous reports on other palladacyclic oligonucleotides.
UV and CD melting studies
Sample preparation for the UV and CD melting studies was identical. Equimolar (1.0 µM) mixtures of
a modified (ON1ao, ON1bo, ON1ao-Pd or ON1bo-Pd) and a natural (ON2a, ON2c, ON2g or ON2t)
oligonucleotide were prepared in 20 mM cacodylate buffer (pH = 7.40), the ionic strength of which
was adjusted to 0.10 M with NaClO4. For the measurement, the samples were placed in quartz
cuvettes with optical path length of 10 mm.
UV melting profiles were acquired by a PerkinElmer Lambda 35 UV/vis spectrophotometer
equipped with a Peltier temperature control unit. Three heating and cooling ramps (10—90 C, 0.5 C
min-1) were carried out on each sample and absorbance at 260 nm recorded at 0.5 C intervals. UV
melting temperatures were determined by Gaussian fitting of the first derivative curves of the UV
melting profiles.
CD spectra were acquired by an Applied Photophysics Chirascan spectropolarimeter equipped
with a Peltier temperature control unit. For each sample, nine spectra at 10 C intervals over a
temperature range of 10—90 C were recorded. At each temperature, the samples were allowed to
equilibrate for either 120 s (unmetallated duplexes) or 1800 s (metallated duplexes) before
measurement.  The wavelength range employed was 220—360 nm. CD melting temperatures were
determined by non-linear least-squares fitting of Equation 1 to plots of molar ellipticity at 285 nm as
a function of temperature.
15N NMR spectrometric studies
Samples for the 15N NMR spectrometric studies were prepared by lyophilizing appropriate amounts of
the aqueous stock solutions of oligonucleotides ON1bo, ON1bo-Pd and ON2t* and redissolving the
residue in D2O, the pD of which was adjusted to 7.8 (20 mM cacodylate buffer) and the ionic strength
21
to 0.10 M (NaClO4). The oligonucleotide concentration of the samples was 150 µM. The spectra were
acquired on a Bruker Biospin 500 MHz instrument equipped with a BBO cryoprobe. For single-stranded
ON1bo, ON1bo-Pd and ON2t*, a single spectrum was recorded at 25 C. With the duplex ON1bo-
PdON2t*, seven spectra were recorded at 10 C intervals over a range of 10—70 C. 22000 scans
spanning 25252 Hz (spectral width) were averaged for a typical spectrum. Default parameters for a 1D
15N NMR measurement were used, with the exception of a longer pre-scan delay (30 µs). The spectra
were processed with an exponential window function, resulting in a line broadening of 2.0 Hz.
Acknowledgments
The help of Dr. Jani Rahkila with 15N NMR spectrometry is gratefully recognized.
References
[1] D. Ukale, T. Lönnberg, ChemBioChem 2021, 22, 1733–1739.
[2] S. Naskar, R. Guha, J. Müller, Angew. Chem. Int. Ed. 2020, 59, 1397–1406.
[3] B. Jash, J. Müller, Chem. Eur. J. 2017, 23, 17166–17178.
[4] Y. Takezawa, J. Müller, M. Shionoya, Chem. Lett. 2017, 46, 622–633.
[5] B. Lippert, P. J. Sanz Miguel, Acc. Chem. Res. 2016, 49, 1537–1545.
[6] P. Scharf, J. Müller, ChemPlusChem 2013, 78, 20–34.
[7] Y. Takezawa, M. Shionoya, Acc. Chem. Res. 2012, 45, 2066–2076.
[8] G. H. Clever, M. Shionoya, Coord. Chem. Rev. 2010, 254, 2391–2402.
[9] T. Nakama, Y. Takezawa, D. Sasaki, M. Shionoya, J. Am. Chem. Soc. 2020, 142, 10153–10162.
[10] Y. Takezawa, L. Hu, T. Nakama, M. Shionoya, M. Shionoya, Angew. Chem. Int. Ed. 2020, 59,
21488–21492.
[11] Y. Takezawa, T. Nakama, M. Shionoya, J. Am. Chem. Soc. 2019, 141, 19342–19350.
[12] J. Kondo, Y. Tada, T. Dairaku, Y. Hattori, H. Saneyoshi, A. Ono, Y. Tanaka, Nat. Chem. 2017, 9,
956–960.
22
[13] A. Ono, H. Kanazawa, H. Ito, M. Goto, K. Nakamura, H. Saneyoshi, J. Kondo, Angew. Chem. Int.
Ed. 2019, 58, 16835–16838.
[14] F. Linares, E. García-Fernández, F. J. López-Garzón, M. Domingo-García, A. Orte, A. Rodríguez-
Díeguez, M. A. Galindo, Chem. Sci. 2019, 10, 1126–1137.
[15] S. Hensel, K. Eckey, P. Scharf, N. Megger, U. Karst, J. Müller, Chem. Eur. J. 2017, 23, 10244–
10248.
[16] T. Ehrenschwender, W. Schmucker, C. Wellner, T. Augenstein, P. Carl, J. Harmer, F. Breher, H.-
A. Wagenknecht, Chem. Eur. J. 2013, 19, 12547–12552.
[17] S. Taherpour, O. Golubev, T. Lönnberg, Inorg. Chim. Acta 2016, 452, 43–49.
[18] B. Jash, J. Müller, Eur. J. Inorg. Chem. 2017, 2017, 3857–3861.
[19] B. Jash, P. Scharf, N. Sandmann, C. Fonseca Guerra, D. A. Megger, J. Müller, Chem. Sci. 2017,
8, 1337–1343.
[20] A. Aro-Heinilä, T. Lönnberg, P. M. Virta, ChemBioChem 2020, 22, 354–358.
[21] X. Guo, F. Seela, Chem. Eur. J. 2017, 23, 11776–11779.
[22] M. Flamme, F. Levi-Acobas, S. Hensel, S. Naskar, P. Röthlisberger, I. Sarac, G. Gasser, J. Müller,
M. Hollenstein, ChemBioChem 2020, 21, 3398–3409.
[23] M. Flamme, P. Rö, F. Levi-Acobas, M. Chawla, R. Oliva, L. Cavallo, G. Gasser, P. Marliè, P.
Herdewijn, M. Hollenstein, ACS Chem. Biol. 2020, 15, 2872–2884.
[24] G. H. Clever, K. Polborn, T. Carell, Angew. Chemie Int. Ed. 2005, 44, 7204–7208.
[25] C. Kaul, M. Müller, M. Wagner, S. Schneider, T. Carell, Nat. Chem. 2011, 3, 794–800.
[26] E.-K. Kim, C. Switzer, ChemBioChem 2013, 14, 2403–2407.
[27] K. Tanaka, M. Shionoya, J. Org. Chem. 1999, 64, 5002–5003.
[28] H. Räisälä, T. Lönnberg, Chem. Eur. J. 2019, 25, 4751–4756.
[29] S. K. Maity, M. A. Hande, T. Lönnberg, ChemBioChem 2020, 21, 2321–2328.
[30] M. Hande, S. Maity, T. Lönnberg, J. Inorg. Biochem. 2021, 222, 111506.
[31] S. Taherpour, H. Lönnberg, T. Lönnberg, Org. Biomol. Chem. 2013, 11, 991–1000.
23
[32] E. Meggers, P. L. Holland, W. B. Tolman, F. E. Romesberg, P. G. Schultz, J. Am. Chem. Soc.
2000, 122, 10714–10715.
[33] P. Scharf, B. Jash, J. A. Kuriappan, M. P. Waller, J. Müller, Chem. Eur. J. 2016, 22, 295–301.
[34] T. Dairaku, K. Furuita, H. Sato, J. Šebera, K. Nakashima, J. Kondo, D. Yamanaka, Y. Kondo, I.
Okamoto, A. Ono, V. Sychrovský, C. Kojima, Y. Tanaka, Chem. Eur. J. 2016, 22, 13028–13031.
[35] Y. Tanaka, S. Oda, H. Yamaguchi, Y. Kondo, C. Kojima, A. Ono, J. Am. Chem. Soc. 2007, 129,
244–245.
[36] O. P. Schmidt, S. Jurt, S. Johannsen, A. Karimi, R. K. O. Sigel, N. W. Luedtke, Nat. Commun.
2019, 10, 4818.
[37] C. B. Reese, Q. Wu, Org. Biomol. Chem. 2003, 1, 3160–3172.
[38] Y. Murata, J. Uenishi, Tetrahedron 2016, 72, 4962–4967.
[39] S. Z. Vatsadze, A. V. Medved’Ko, S. A. Kurzeev, O. I. Pokrovskiy, O. O. Parenago, M. O.
Kostenko, I. V. Ananyev, K. A. Lyssenko, D. A. Lemenovsky, G. M. Kazankov, V. V. Lunin,
Organometallics 2017, 36, 3068–3075.
[40] S. K. Maity, T. Lönnberg, Chem. Eur. J. 2018, 24, 1274–1277.
[41] S. K. Maity, T. A. Lönnberg, ACS Omega 2019, 4, 18803–18808.
[42] R. B. Martin, Acc. Chem. Res. 1985, 18, 32–38.
[43] H. M. Bdour, J. L. F. Kao, J. S. Taylor, J. Org. Chem. 2006, 71, 1640–1646.
[44] A. Stukalov, V. V. Suslonov, M. A. Kuznetsov, Eur. J. Org. Chem. 2018, 2018, 1634–1645.
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Entry for the Table of Contents
Oligodeoxynucleotides incorporating either an α or a β anomer of an 15N-labelled palladacyclic
acetophenone oxime C-nucleoside were synthesized and hybridized with various natural
counterparts. UV, CD and 15N NMR data were consistent with a stable Pd(II)-mediated base pair in
the middle of the duplex but failed to prove it conclusively.
Institute and/or researcher Twitter usernames: @UniTurku