Transporting excess electrons along potential energy gradients provided by 2 ' -deoxyuridine derivatives in dna

Article abstract of DOI:10.1002/anie.201202141

LUMO-level dependent: Chemically modified DNA molecules containing 2'-deoxyuridine (dU) derivatives with various LUMO energy levels have been synthesized to manipulate electron-transfer efficiencies. By arranging thymidine, the dU derivatives, and 5-fluor

Full text of DOI:10.1002/anie.201202141

Angewandte
Chemie
DOI: 10.1002/anie.201202141
Electron Transport in DNA
Transporting Excess Electrons along Potential Energy Gradients
Provided by 2’-Deoxyuridine Derivatives in DNA**
Takeo Ito,* Yuta Hamaguchi, Kazuhito Tanabe, Hisatsugu Yamada, and Sei-ichi Nishimoto*
Charge transfer through peptides and proteins is one of the
most important reactions in the processes of photosynthesis,
signal transduction, respiration, and some enzymatic activ-
ities.[1–5] DNA duplexes can also transport holes and excess
electrons over a distance,[3–23] and such charge-transfer
reactions in DNA might be involved in the recognition of
damaged DNA bases by DNA repair enzymes.[24] In addition,
it has been demonstrated that electron-transporting DNA
could be used as a device for genotyping and for single-
nucleotide polymorphism analysis.[25–29]
use modified DNA analogues that overcome the aforemen-
tioned shortcomings of natural DNA bases.
In this study, we developed DNA containing uracil (U)
derivatives with different LUMO energy levels, and examined
the regulation and directional control of EET in DNA. Our
temporal goal is to construct molecular diode-like DNA
nanostructures[32–34] in which the direction and efficiency of
EET could be arbitrarily controlled depending on the
chemical structures of the intervening DNA bases. We
investigated photoinduced electron transport from the
DNA-tethered photoinduced electron donor phenothiazine
(PTZ; Eox* = ꢀ2.7 V vs SCE)[35] to the co-inserted 5-bro-
mouracil (BrU) through the intervening U derivatives. Product
analysis clearly showed that injected electrons migrated
according to the potential energy gradient of the LUMOs of
U derivatives, which is, to the best of our knowledge, the first
example of the manipulation of the direction of EET using
DNA analogues.
As candidates for replacing T in DNA as excess electron
carriers, we chose four 2’-deoxyuridine derivatives: 2’-deoxy-
uridine (dU), 5-fluoro-2’-deoxyuridine (dFU), 5-hydroxy-2’-
deoxyuridine (dOHU), and 2’-deoxypseudouridine (dPU). Our
preliminary density functional theory calculation (B3LYP/6-
31G*) suggested that the LUMO levels of the derivatives are
sufficiently high for transporting excess electrons: LUMO
level of dT, ꢀ1.18 eV; of dU, ꢀ1.28 eV; of dFU, ꢀ1.53 eV; of
dPU, ꢀ1.39 eV and of dOHU, ꢀ1.36 eV (Figure 1). Also, the
electron affinities (EAs) of T, U, and 5-fluorouracil (FU) have
been previously reported as 1.56 eV (T), 1.62 eV (U), and
1.82 eV (FU), respectively.[36]
To date, studies on hole transfer (HT) through the highest
occupied molecular orbital (HOMO) of DNA bases have
demonstrated that holes on DNA migrate over long distances
mainly between guanine–cytosine (G-C) base pairs and
partially between adenine–thymine (A-T) base pairs.
Recently, highly efficient HT was achieved by replacing the
A-T base pair with the 7-deazaguanine–T base pair, which has
a higher HOMO energy level than the A-T base pair.[13]
Electrons injected into DNA also migrate along the
duplex through the lowest unoccupied molecular orbital
(LUMO), most likely between C and T by means of
a thermally activated hopping mechanism at ambient temper-
ature.[17] In contrast with the HT in DNA, the efficiency of
excess electron transfer (EET) from a photoinduced electron
donor over a distance through DNA bases has been reported
as being low. This is partly explained by the fast charge
recombination between the DNA-tethered electron donor
and the excess electron, and kinetically competitive proton
transfer between the radical anion of C (CꢀC) and its
complementary G.[30,31] If one considers the redox stability
of DNA bases, nanoscale electronic devices based on EET
chemistry are seemingly preferable, because HT in DNA
results in the oxidation of G. Although many issues remain
regarding the durability of redox chemical reactions, one of
the strategies for developing novel DNA-based devices is to
Figure 1. Chemical structures of thymine (T), uracil (U), 5-fluorouracil
(FU), 5-hydroxyuracil (OHU), 5-bromouracil (BrU), and pseudouracil
(PU).
[*] Dr. T. Ito, M. Sc. Y. Hamaguchi, Dr. K. Tanabe, Dr. H. Yamada,
Prof. Dr. S. Nishimoto
Department of Energy and Hydrocarbon Chemistry
Graduate School of Engineering, Kyoto University
Kyoto Daigaku Katsura, Nishikyo-ku, Kyoto 615-8510 (Japan)
E-mail: takeoit@scl.kyoto-u.ac.jp
EET efficiency was investigated by product analysis using
polyacrylamide gel electrophoresis (PAGE). PTZ was placed
in the duplex DNA by conventional phosphoramidite
chemistry, and BrU was used as a chemical probe for detecting
excess electrons that migrated from PTZ (Figure 2).[18] Once
an electron is captured by BrU, the spontaneous (k ꢁ 109 sꢀ1
for isolated BrU)[37] release of a bromide anion yields the
corresponding uracil-5-yl radical, which in turn abstracts one
hydrogen from the 5’-adjacent deoxyribose. The sugar radical
niqda894@kitc.city.kyoto.lg.jp
[**] This work was supported by a Grant-in-Aid for the Global COE
program “International Center for Integrated Research and
Advanced Education in Materials Science” from the Ministry of
Education, Culture, Sports, Science and Technology (MEXT) of the
Japanese Government. A part of this study was performed with the
support of the Radioisotope Research Center of Kyoto University.
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/anie.201202141.
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might be the difference in the LUMO distribution between
PU and the other U derivatives.
In addition, the durability of the U radical anions was
confirmed by checking stability during the radiation-induced
reduction of the derivatives in aqueous solution, because
irreversible trapping of excess electrons results in the
retardation of long-range electron transfer. One-electron
reduction of T, U, FU, BrU, and OHU by hydrated electrons
(eaqꢀ) generated as a consequence of water radiolysis under
oxygen-free conditions was examined, and decomposition of
U derivatives was monitored using HPLC. However, no
remarkable decomposition was observed, except in the cases
of BrU and OHU (Figure S3 in the Supporting Information).
Considering the chemical reactivities of the U derivatives, we
chose T, U, and FU as electron-transporting molecules for
constructing artificial DNA. EET efficiency through the
consecutive U bases increased (F-ODN1 > U-ODN1 > T-
ODN1) as the LUMO energy level of the uridine bases
decreased (T> U > FU).
Figure 2. Chemical structure of phenothiazine (PTZ). Excess electrons
injected from photoexcited PTZ (PTZ*) migrate through T and U
derivatives and react irreversibly with BrU or recombine with the PTZ
radical cation (PTZ+·).
eventually affords alkali-labile products as a result of
reactions with water.[38–40] Thus, the excess electrons that
migrated from PTZ to BrU could be evaluated by quantifying
the amount of strand-cleavage products obtained after
piperidine-catalyzed hydrolysis of the photoexposed DNA.
To evaluate the relative electron-transfer efficiency
through the U derivatives, the photoinduced electron-transfer
reaction was investigated in DNA containing four consecutive
U derivatives between PTZ and BrU (PTZ-ODN1/X-ODN1;
Figure 3a). DNAwas exposed to UV light at 365 nm under an
Encouraged by these results, we next prepared a DNA
containing PTZ in the middle of the duplex, four consecutive
U derivatives on both the 3’ and 5’ sides of PTZ, and two Br
U
bases as electron-transfer probes. For this experiment,
enantiomeric (R)- and (S)-PTZ phosphoramidites were
synthesized and inserted into the duplex to evaluate the
effect of the localization of PTZ in DNA (Figure 4a). As
summarized in Figure 4b, the electron-transfer efficiencies
through the U bases in the 5’!3’ direction (through the YYY
sequence) and in the 3’!5’ direction (through the ZZZ
sequence) were compared. When Y and Z were identical,
electron migration in the 3’!5’ direction was predominant (in
the case of TT-ODN2, the yields of the electron-transfer
Figure 3. EET from photoexcited PTZ to BrU through four uracil
derivatives (X). a) DNA sequences of PTZ-ODN1 and X-ODN1; b) 32P-
&
*
radiolabeled duplex DNA (PTZ-ODN1/X-ODN1; X=( ) T, (ꢀ) U, ( )
F
OH
P
~
*
U, ( ) U, ( ) U) in buffer solution (10 mm phosphate, 90 mm
NaCl, pH 7.0) was exposed to UV light (365 nm) for the indicated
periods at 48C, followed by treatment with piperidine at 908C for
30 min. DNA fragments that corresponded to electron-transfer prod-
ucts were quantified using PAGE.
N2 atmosphere and then treated with piperidine at 908C.
Uracil derivatives, including BrU, do not absorb UV light at
wavelengths above 350 nm (Figure S1 in the Supporting
Information). PAGE of the products revealed the formation
of electron-transfer products with a yield that varied depend-
ing on the sequences of the inserted U bases (Figure S2 in the
Supporting Information). The electron-transfer efficiency was
remarkably higher in the FU- or U-containing duplexes (PTZ-
ODN1/F-ODN1 and PTZ-ODN1/U-ODN1) than it was in
the other duplexes (Figure 3b). The thermal stability of the
duplexes, as evaluated by melting temperature (Tm), suggests
Figure 4. Asymmetrical EET from photoexcited PTZ to BrU through
uracil derivatives. a) Chemical structure of (R)- and (S)-PTZ inserted in
the middle of oligodeoxynucleotides; b) 32P-radiolabeled duplex DNA
((R)-PTZ-ODN2/YZ-ODN2; Y, Z=T, U, U) in buffer solution (10 mm
phosphate, 90 mm NaCl, pH 7.0) was exposed to UV light (365 nm) at
48C, followed by piperidine treatment.
P
that U and OHU slightly destabilize the duplex (Table S1 in
the Supporting Information). Therefore, local structural
distortion at the PU or OHU stretches should diminish the
apparent electron-transfer efficiency.[35] Another reason for
the inefficient electron transfer in the PU-containing duplexes
F
2
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Figure 5. EET through various sequences of uracil derivatives. a) DNA sequences used in this experiment; b) sequence-dependent EET efficiencies
as determined from the yield of DNA-cleavage products. The sequence in the gray box was changed as indicated in the diagram, which depicts
the calculated LUMO levels of uracil derivatives and PTZ*.
products were too low for the efficiencies to be compared),
which is in accordance with the results of previous
reports.[31,41] The intrinsic electron flow could be reversed by
inserting U derivatives, that is, electrons moved to U
derivatives with higher EA. For example, electron transfer
in the 3’!5’ direction was predominant in the TU-ODN1-
containing duplex, but migration in the other direction
became predominant when the sequence was replaced with
UT-ODN1. Asymmetrical localization of PTZ in the
duplexes seemed less likely, because the enantiomeric (R)-
and (S)-PTZ-containing ODNs showed essentially the same
electron-transport properties (Figure S4 in the Supporting
Information). Nonlocalization of the photoinduced electron
donor has also been confirmed in the case of diaminonaph-
thalene-tethered DNA duplexes.[31]
Figure 5b, excess electron hopping through U stretches in
which the LUMO energy level is flat (DS 1, DS 7, and DS 11)
or upstream from the 3’!5’ direction (DS 6, DS 9, and DS 10)
was apparently inefficient compared with that observed
through DS 2–DS 5 and DS 8, probably because of fast
charge recombination. It was also apparent that the multistep
energy gradient enhanced the charge separation, as observed
for DS 4 and DS 5. The role of the nucleobase at the electron-
injection moiety seems to be important in preventing initial
charge recombination,[44] because insertion of FU at the 3’-
adjacent base of PTZ apparently lowered the electron-
transfer efficiency compared with the case of PTZ-ODN1/
F-ODN1. Back and forth EET between intervening U bases
with a flat energy level may enhance the lifetime of excess
electrons on the duplex and result in back electron transfer to
PTZ+C. Interaction between adjacent DNA bases could alter
their LUMO levels to some extent; nevertheless, the current
study demonstrates that it is possible to estimate electron-
transfer efficiency based on the LUMO levels of the isolated
uracil derivatives.
In conclusion, we have synthesized artificial DNA
duplexes containing U derivatives as alternative pyrimidine
bases to T to modulate the electron-transport properties of
DNA. Our successful control of the directionality of the
electron transfer by using T, U, and FU may widen the
potential applications of artificial DNA as a novel electronic
device; for example, the control of electron transfer on
multidimensional DNA duplexes might be possible by using
branched duplexes containing modified DNA bases.
Mechanistic investigation of DNA-mediated charge trans-
fer has revealed that excess electrons hop between nucleo-
bases, most likely between pyrimidine bases. Moreover, it has
been suggested that back electron transfer to the radical
cation of the electron donor occurs rapidly and effi-
ciently.[42–45] Thus, the remarkable enhancement of the
apparent electron-transfer efficiency observed in the cases
F
of U- or U-containing DNA could be explained by retarda-
tion of the back electron-transfer process; in other words, an
electron injected from PTZ* to the adjacent Teither migrates
further to the next U bases or recombines with PTZ+·.
Therefore, the LUMO energy gap between the adjacent Tand
the next consecutive U derivatives should affect the relative
yield of back electron transfer. It is not clear why electron
transfer from 3’ to 5’ of pyrimidine bases is preferable, but it
has been suggested that asymmetrical orbital overlap around
the electron-deficient nucleobase intermediate may affect the
direction of HT in DNA.[46] Such directionality should be
considered if we develop DNA devices for genotyping based
on the charge-transfer properties of DNA.
Finally, we explored the construction of DNA containing
U derivatives to produce various LUMO energy potential
gradients along the strands (Figure 5). In this experiment, the
PTZ-adjacent nucleobase was changed (T-, U-, and F-PTZ-
ODN3) and three U derivatives were inserted on the opposite
strands (TTT-, UUU-, FFF-, TUF-, and UFF-ODN3). Eleven
duplex DNAs (DS 1–11; Figure 5b) obtained by the combi-
nation of the oligodeoxynucleotides were used. As depicted in
Received: March 18, 2012
Published online: && &&, &&&&
Keywords: DNA · electron transport · photochemistry ·
.
radical ions · radical reactions
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Communications
Electron Transport in DNA
T. Ito,* Y. Hamaguchi, K. Tanabe,
H. Yamada,
S. Nishimoto*
&&&& — &&&&
Transporting Excess Electrons along
Potential Energy Gradients Provided by
2’-Deoxyuridine Derivatives in DNA
LUMO-level dependent: Chemically
modified DNA molecules containing 2’-
deoxyuridine (dU) derivatives with vari-
ous LUMO energy levels have been
synthesized to manipulate electron-
transfer efficiencies. By arranging thymi-
dine, the dU derivatives, and 5-fluoro-2’-
deoxyuridine in order of their LUMO
levels, the efficiency and the directionality
of photoinduced electron transport in
DNA could be regulated.
Angew. Chem. Int. Ed. 2012, 51, 1 – 5
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These are not the final page numbers!