One-Electron Attachment Reaction of B- and Z-DNA Modified by 8-Bromo-2′-deoxyguanosine Takumi Kimura, Kiyohiko Kawai, Sachiko Tojo, and Tetsuro Majima* The Institute of Scientific and Industrial Research (SANKEN), Osaka University, Mihogaoka 8-1, Ibaraki, Osaka 567-0047, Japan
[email protected] Received August 15, 2003
The one-electron attachment reaction of 8-bromo-2′-deoxyguanosine (BrG) in DNA was studied by comparing that in B- and Z-DNA. Oligodeoxynucleotides (ODNs) modified by BrG were synthesized as Z-DNA in which the syn-conformation deoxyguanosine is stabilized by steric interference between the 8-bromo group of BrG and the sugar moiety. Debromination from the BrG-modified ODNs occurred from the one-electron attachment during the γ-radiolysis. The structural dependence of B- and Z-DNA was observed for the one-electron attachment reaction. The conversion of BrG was higher in Z-DNA than in B-DNA. Because the solvent-accessible surface of the purine base in Z-DNA is greater than that in B-DNA, it is demonstrated that the reactivity of purine base C8 is enhanced in Z-DNA compared to that in B-DNA. Introduction DNA local conformations are considered to play an important role in biological phenomena such as DNAprotein interactions.1,2 The existence of Z-DNA in living cells has been suggested by observation of Z-DNA stabilization by negative supercoil3 and prediction of Z-DNA formation in sequences of human genes,4 since the lefthanded Z-DNA was discovered by Poul and Jovin in 1972.5 (CG)n repeats have the highest potential to form Z-DNA, which has a left-handed zigzag backbone and unusual syn-conformation of purine base. However, the biological role of Z-DNA has not been determined. Because the Z-DNA stabilization by the negative supercoil is relaxed by an enzyme topoisomerase, Z-DNA is considered to be a temporal structure. In 1999, Rich et al. reported the X-ray crystallography of double-stranded RNA adenosine deaminase (ADAR1) possessing the potential to undergo Z-DNA binding.6 This result suggested Z-DNA effects the gene expression.7,8 Therefore, the biological role of Z-DNA has become a topical research subject of Z-DNA.9,10 Dinucleotide repeats have the potential to form alternative DNA structures such as Z-DNA, H-DNA, and cruciform * Author to whom correspondence should be addressed. Phone: Japan +6-6879-8495. Fax: Japan +6-6879-8499. (1) Cozzarelli, N. R.; Wang, J. C. DNA Topology and Its Biological Effects; Cold Spring Harbor Laboratory Press: Cold Spring Harbor, NY, 1990. (2) Travers, A. A. Annu. Rev Biochem. 1989, 58, 427-435. (3) Liu, L. F.; Wang, J. C. Proc. Natl. Acad. Sci. U.S.A. 1987, 84, 7024-7027. (4) Schroth, G. P.; Chou, P. J.; Ho, P. S. J. Biol. Chem. 1992, 267, 11846-11855. (5) Pohl, F. M.; Jovin, T. M. J. Mol. Biol. 1972, 67, 375-396. (6) Schwaltz, T.; Rould, M. A.; Lowenhaupt, K. H.; Rich, A. Science 1999, 284, 1841-1845. (7) Tiesman, J.; Rizzino, A. Gene 1990, 96, 311-312. (8) Vlach, J.; Dvorak, M.; Bartunek, P.; Pecenka, V.; Travnicek, M.; Sponar, J. Biochem. Biophys. Res. Commun. 1989, 158, 737-742.
DNA.11,12 Because of the structural and sequential polymorphic nature of the dinucleotide repeats, it is suggested that the Z-DNA-forming sequences may provide a source of genetic variation if they occur in regions that are important for the regulation of gene activities. To investigate the biological role of Z-DNA, the development of selective Z-DNA detection methods is desired. Clarification of the characteristic Z-DNA reactivity at the oligomer level is necessary for the development of a Z-DNA probe. Although the Z conformation is not adopted in unmodified ODNs under physiological conditions, Rich previously reported that the bromination of poly d(CG), giving 5-bromo-2′-deoxycytosine or BrG, readily stabilizes the Z-DNA.13 The presence of a bulky substituent such as the bromine atom or phenyl group14 at the C8 position of guanosine destabilizes the normal anti orientation of the base, sterically constraining the molecule to the syn conformation. By incorporating 8-methylguanosine, Sugiyama et al. synthesized the 5-halouracil-modified Z-form DNA and demonstrated the potential of the 5-halouracil photoreaction as a probe for Z-DNA, in which a Zconformation specific photoproduct offers the detection of Z-DNA in vivo.15-17 More recently, Eritja demonstrated (9) Schade, M.; Turner, C. J.; Kuhne, R.; Schmeieder, P.; Lowenhaupt, K.; Herbert, A.; Rich, A.; Oschkinat, H. Proc. Natl. Acad. Sci. U.S.A. 1999, 96, 12465-12470. (10) Schwartz, T.; Behike, J.; Lowenhaupt, K.; Heinemann, U.; Rich, A. Nat. Struct. Biol. 2001, 8, 761-765. (11) Wells, R. D. J. Biol. Chem. 1988, 263, 1095-1098. (12) Rich, A. Gene 1993, 135, 99-109. (13) Mo¨ller, A.; Nordheim, A.; Kozlowski, S. A.; Patel, D. J.; Rich, A. Biochemistry 1984, 23, 54-62. (14) Gannett, P. M.; Heavner, S.; Daft, J. R.; Shaughnessy, K. H.; Epperson, J. D.; Greenbaum, N. L. Chem. Res. Toxicol. 2003, 16, 13851394. (15) Kawai, K.; Saito, I.; Sugiyama, H. J. Am. Chem. Soc. 1999, 121, 1391-1392. (16) Kawai, K.; Saito, I.; Kawashima, E.; Ishido, Y.; Sugiyama, H. Tetrahedron Lett. 1999, 40, 2589-2592.
10.1021/jo035194q CCC: $27.50 © 2004 American Chemical Society
Published on Web 01/21/2004
J. Org. Chem. 2004, 69, 1169-1173
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the Z-DNA stabilization of decamers containing alternating CG sequences in which two of the five residues were replaced by BrG.18 However, the Z-DNA stabilization is limited only in a few decamers. Mulazzani et al. investigated the reactions of a hydrated electron (eaq-) with 8-bromoguanosine during γ-radiolysis of 8-bromoguanosine in aqueous solution based on the product analysis.19 Because guanosine was detected as a single product, it is possible to analyze Z-DNA-specific reactions during the γ-radiolysis of the BrG-modified ODNs. Here, we quantified the debromination yield of various BrG-modified B-form and Z-form ODNs in the one-electron attachment reaction during the γ-ray irradiation. On the basis of the experimental results, the one-electron attachment reaction is discussed with respect to a specific probe for the existence of a local Z-DNA in living cell.
TABLE 1. Sequence of
BrG-modified ODNs and Concentrations of Sodium Chloride and Magnesium Chloride at the Midpoint of the B-Z Transitiona
1 2 3 4 5 6 7 8 9
ODNs
NaCl (mM)
MgCl2 (mM)
d(CGCGCG)2 d(CGCm8GCG)2 d(CGCBrGCG)2 d(CGCGCGCG)2 d(CBrGCGCGCG)2 d(CGCBrGCGCG)2 d(CGTGCACG)2 d(CBrGTGCACG)2 d(CGTBrGCACG)2
2600
