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T. Ito et al. / Bioorg. Med. Chem. Lett. 17 (2007) 6129–6133
be competing with the electron migration along the
strand, and thus lowering the reactivity of Tg. It is also
expected that reduction of guanine is less efficient be-
tance until they are trapped by cytosine or Tg in the se-
quences, on the other hand, guanine could diminish the
efficiency of electron attachment to the strands because
of its low electron affinity. Our fluorescence quenching
experiment showed that electron affinity of dTg is lower
than that of dT, and thus which affects reductive elec-
tron migration in DNA. Electron affinity of DNA le-
sions is possibly important to understand DNA
modification mechanisms involved in the early stages
of DNA damage formation by ionizing radiation and
UV-light.
cause of the high reduction potential (E < ꢀ3.00 V
red
1
2
0
0
vs SCE), especially in the sequence of 5 -GGTgG-3 ,
which is well consistent with the reactivity obtained in
4
a
this study. As discussed in our previous report, 6-hy-
droxy-5,6-dihydrothymin-5-yl radical generated from
Tg could be further reduced by adjacent guanines due
to high electron affinity of the radical, as a result, the
photolysis may yield guanine modifications in the se-
quence. In addition, it has been predicted that the 5-yl
radical adds to the adjacent guanines to form cross-
1
3,14
linked products.
photolysis of Tg-containing oligodeoxynucleotide pos-
For that reason, we attempted a
Acknowledgment
0
0
sessing multiple guanines (5 -CTTGGGTgGCT-3 ) in
the presence of flavins. Sodium dithionite or EDTA
was employed for reducing FAD into FADH in the
This work was partially supported by Grants-in-Aid for
Young Scientists (B) from the Ministry of Education,
Culture, Sports, Science, and Technology of Japan.
ꢀ
present reaction. After piperidine-catalyzed hydrolysis,
the products were analyzed by gel electrophoresis, but
no alkaline-labile products of guanine were detected
Supplementary data
(
(
Supplementary data). Photoreduction by phenothiazine
4,15
*
PTZ, E = ꢀ2.7 V),
an alternative electron donor
ox
for avoiding the possibility of reduction of the transient
guanine radical cation by excess dithionite, did not give
any notable products. Consistent with the results with
the G-rich 4-mer long oligodeoxynucleotides, repair of
Tg was not observed in the longer sequence, which sug-
gests that the G-rich oligodeoxynucleotides are intrinsi-
cally less reactive toward photoexcited reduced forms of
the flavins.
References and notes
. Fukuzumi, S.; Tanaka, T. In Photoinduced Electron
Transfer Part C (Photoinduced Electron Transfer Reac-
tions: Organic Substrates); Fox, M. A., Chanon, M., Eds.;
Elsevier: Amsterdam, 1998; pp 636–687, and the references
therein.
. (a) Jorns, M. S. J. Am. Chem. Soc. 1987, 109, 3133; (b)
Jorns, M. S.; Wang, B.; Jordan, S. P.; Chanderkar, L. P.
Biochemistry 1990, 29, 552; (c) Sancar, A. Chem. Rev.
1
With regard to interaction between electrons and DNA
containing modified nucleic acid bases, we have demon-
strated that Tg in DNA sequences does not prevent reduc-
2
4
a
tive electron migration along the duplex. To compare
electron affinities between dTg and dT by reductive fluo-
rescence quenching analysis, 1-aminopyrene (AP,
2
003, 103, 2203.
3
. (a) Song, P. J. Am. Chem. Soc 1969, 91, 1850; (b) Barrio, J.
R.; Tolman, G. L.; Leonard, N. J.; Spencer, R. D.; Weber,
G. Proc. Natl. Acad. Sci. U.S.A. 1973, 70, 941; (c) Visser,
A. J. W. G. Photochem. Photobiol. 1984, 40, 703; (d)
Chosrowjan, H.; Taniguchi, S.; Mataga, N.; Tanaka, F.;
Visser, A. J. W. G. Chem. Phys. Lett. 2003, 378, 354; (e)
Islam, S. D. M.; Susdorf, T.; Penzkofer, A.; Hegemann, P.
Chem. Phys. 2003, 295, 137; (f) Walsh, J. D.; Miller, A.-F.
J. Mol. Struct. Theochem. 2003, 623, 185.
*
Eox = ꢀ2.6 V) in 50% acetonitrile aqueous solution was
employed as a photoinduced electron donor. Upon exci-
tation of AP at 365 nm, fluorescence intensity
1
6
(
kmax = 443 nm, s = 5.2 ns) decreased as increasing
0
the amount of dT, and the Stern–Volmer plot gave a
quenching rate constant of
k = (9.8 ± 0.5) ·
(Supplementary information). On the other
slightly lower quenching rate constant
q
1
0
ꢀ1 ꢀ1
M s
1
0
hand,
a
9
ꢀ1 ꢀ1
4. (a) Ito, T.; Kondo, A.; Terada, S.; Nishimoto, S. J. Am.
Chem. Soc. 2006, 128, 10934; (b) Ide, H.; Otsuki, N.;
Nishimoto, S.; Kagiya, T. J. Chem. Soc. Perkin Trans. 2
[
k = (3.4 ± 0.1) · 10 M
q
s ] was obtained for dTg
suggesting that reduction potential of Tg is more negative
than that of dT. The lower electron affinity of Tg might be
the reason for the efficient electron migration through
1
985, 1387; (c) Nishimoto, S.; Ide, H.; Otsuki, N.;
Nakamichi, K.; Kagiya, T. J. Chem. Soc. Perkin Trans.
2 1985, 1127.
4
a
Tg-containing oligodeoxynucleotides.
5
. (a) Iida, S.; Hayatsu, H. Biochim. Biophys. Acta 1971, 228,
1; (b) Vaishnav, Y.; Holwitt, E.; Swenberg, C.; Lee, H. C.;
Kan, L. S. J. Biomol. Struct. Dyn. 1991, 8, 935.
. Monoexponential fluorescence decay lifetime of FMN
In conclusion, photoinduced reduction of dTg and Tg-
containing oligodeoxynucleotides sensitized by flavins
was investigated by quantitative product analysis and
fluorescence quenching. Although oxidized form of
FAD may discriminate the structural difference between
dTg and dT with relatively weak interactions, this did
not affect photoinduced reductive repair of Tg into dT.
As demonstrated above and in our previous attempts,
6
(
s = 4.70 ns), or longer component of biexponential decay
lifetime of FAD (s = 2.82 ns, Ref. 3c) was employed for
the Stern–Volmer analysis.
7
. (a) Clark, J. M.; Pattabiraman, N.; Jarvis, W.; Beardsley,
J. P. Biochemistry 1987, 26, 5404; (b) Basu, A. K.;
Loechler, E. L.; Leadon, S. A.; Essigmann, J. M. Proc.
Natl. Acad. Sci. U.S.A. 1989, 86, 7677; (c) Mlasklewicz,
K.; Miller, J.; Ornstein, R.; Osman, R. Biopolymers 1995,
35, 113; (d) Iwai, S. Chem. Eur. J. 2001, 7, 4344.
ꢀ
ꢀ
both FADH and FMNH can induce repair of Tg in
single-stranded DNA in a sequence-dependent manner.
We anticipate that electrons could migrate a short dis-