Abstract
Nicks and flaps are intermediates in various processes of DNA metabolism, including replication and repair. Photoaffinity modification was employed in studying the interaction of the replication protein A (RPA) and flap endonuclease 1 (FEN-1) with DNA duplexes similar to structures arising during long-patch base excision repair. The proteins were also tested for effect on DNA polymerase β (Polβ) interaction with DNA. Using Polβ, a photoreactive dTTP analog was added to the 3" end of an oligonucleotide flanking a nick or a flap in DNA intermediates. The character and intensity of protein labeling depended on the type of intermediates and on the presence of the phosphate or tetrahydrofuran at the 5" end of a nick or a flap. Photoaffinity labeling of Polβ substantially (up to three times) increased in the presence of RPA or FEN-1. Various DNA substrates were used to study the effects of RPA and FEN-1 on Polβ-mediated DNA synthesis with displacement of a downstream primer. In contrast to FEN-1, RPA had no effect on DNA repair synthesis by Polβ during long-patch base excision repair.
Similar content being viewed by others
REFERENCES
Biade, S., Sobol, R.W., Wilson, S.H., and Matsumoto, Y., J. Biol. Chem., 1998, vol. 273, pp. 898–902.
Dianov, G.L., Prasad, R., Wilson, S.H., and Bohr, V.A., J. Biol. Chem., 1999, vol. 274, pp. 13741–13743.
Fortini, P., Parlanti, E., Sidorkina, O.M., et al., J. Biol. Chem., 1999, vol. 274, pp. 15230–15236.
Frosina, G., Fortini, P., Rossi, O., et al., J. Biol. Chem., 1996, vol. 271, pp. 9573–9578.
Klungland, A. and Lindahl, T., EMBO J., 1997, vol. 16, pp. 3341–3348.
Prasad, R., Dianov, G.L., Bohr, V.A., and Wilson, S.H., J. Biol. Chem., 2000, vol. 275, pp. 4460–4466.
Matsumoto, Y., Kim, K., and Bogenhagen, D.F., Mol. Cell. Biol., 1994, vol. 14, pp. 6187–6197.
Kelman, Z., Oncogene, 1997, vol. 14, pp. 629–640.
Podlutsky, A.J., Dianova, I.I., Podust, V.N., et al., EMBO J., 2001, vol. 20, pp. 1477–1482.
Prasad, R., Lavrik, O.I., Kedar, P., et al., J. Biol. Chem., 2001, vol. 276, pp. 32411–32414.
Collins, K.L. and Kelly, T.J., Mol. Cell. Biol., 1991, vol. 11, pp. 2108–2115.
DeMott, M.S., Zigman, S., and Bambara, R.A., J. Biol. Chem., 1998, vol. 273, pp. 27 492–27 498.
Podust, V.N. and Hubscher, U., Nucleic Acids Res., 1993, vol. 21, pp. 841–846.
Khlimankov, D.Yu., Rechkunova, N.I., Kolpashchikov, D.M., et al., Mol. Biol., 2001, vol. 35, pp. 827–835.
Lavrik, O.I., Nasheuer, H.-P., Weisshart, K., et al., Nucleic Acids Res., 1998, vol. 26, pp. 602–607.
Lavrik, O.I., Kolpashchikov, D.M., Nasheuer, H.-P., et al., FEBS Lett., 1998, vol. 441, pp. 186–190.
Kolpashchikov, D.M., Weisshart, K., Nasheuer, H.-P., et al., FEBS Lett., 1999, vol. 450, pp. 131–134.
Lavrik, O.I., Kolpashchikov, D.M., Weisshart, K., et al., Nucleic Acids Res., 1999, vol. 27, pp. 4235–4240.
Kolpashchikov, D.M., Khodyreva, S.N., and Lavrik, O.I., Dokl. Akad. Nauk, 2000, vol. 372, pp. 824–826.
Kolpashchikov, D.M., Khodyreva, S.N., Khlimankov, D.Yu., et al., Nucleic Acids Res., 2001, vol. 29, pp. 373–379.
Beard, W.A. and Wilson, S.H., Methods Enzymol., 1995, vol. 262, pp. 98–107.
Henricksen, L.A., Umbricht, C.B., and Wold, M.S., J. Biol. Chem., 1994, vol. 269, pp. 11121–11132.
Kolpashchikov, D.M., Zakharenko, A.L., Dezhurov, S.V., et al., Bioorgan. Khim., 1999, vol. 25, pp. 129–136.
Sambrook, J., Fritsch, E.F., and Maniatis, T., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor, N.Y.: Cold Spring Harbor Lab. Press, 1989.
Yamshchikov, V.F., Metody molekulyarnoi genetiki i gennoi inzhenerii (Methods in Molecular Genetics and Gene Engineering), Salganik, R.I., Ed., Novosibirsk: Nauka, 1990, pp. 145–154.
Laemmli, U.K., Nature, 1970, vol. 227, pp. 680–685.
Wilson, S.H., Mutat. Res., 1998, vol. 407, pp. 203–215.
Srivastava, D.K., Berg, B.J., Prasad, R., et al., J. Biol. Chem., 1998, vol. 273, pp. 21 203–21 209.
Slupphaug, G., Eftedal, I., Kavli, B., et al., Biochemistry, 1995, vol. 34, pp. 128–138.
Lao, Y., Gomes, X.V., Ren, Y., et al., Biochemistry, 2000, vol. 39, pp. 850–859.
Khlimankov, D.Yu., Petruseva, I.O., Rechkunova, N.I., et al., Bioorgan. Khim., 2001, vol. 27, pp. 205–209.
Harrington, J.J. and Lieber, M.R., EMBO J., 1994, vol. 13, pp. 1235–1246.
Ranalli, T.A., DeMott, M.S., and Bambara, R.A., J. Biol. Chem., 2002, vol. 277, pp. 1219–1227.
Maga, G., Frouin, I., Spadari, S., and Hubscher, U., J. Biol. Chem., 2001, vol. 276, pp. 18235–18242.
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Khlimankov, D.Y., Rechkunova, N.I., Khodyreva, S.N. et al. Interaction of Replication Protein A and Flap Endonuclease 1 with DNA Duplexes Containing a Nick or a Flap. Molecular Biology 36, 849–856 (2002). https://doi.org/10.1023/A:1021642228046
Issue Date:
DOI: https://doi.org/10.1023/A:1021642228046