A helical arch allowing single-stranded DNA to thread through T5 5'-exonuclease

Abstract

THE 5'-exonucleases are enzymes that are essential for DNA replication and repair1. As well as their exonucleolytic action, removing nucleotides from the 5'-end of nucleic acid molecules such as Okazaki fragments2, many 5'-3'-exonucleases have been shown to possess endonucleolytic activities3,4. T5 5'-3'-exonuclease shares many similarities with the amino termini of eubacterial DNA polymerases5, although, unlike eubacteria, phages such as T5, T4 and T7 express polymerase and 5'-exonuclease proteins from separate genes. Here we report the 2.5-Å crystal structure of the phage T5 5'-exonuclease, which reveals a helical arch for binding DNA. We propose a model consistent with a threading mechanism in which single-stranded DNA could slide through the arch, which is formed by two helices, one containing positively charged, and the other hydrophobic, residues. The active site is at the base of the arch, and contains two metal-binding sites.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1

    Kornberg, A. & Baker, T. DNA Replication 2nd edn (W. H. Freeman, New York, 1992).

  2. 2

    Okazaki, R., Arisawa, M. & Sugino, A. Proc. natn. Acad. Sci. U.S.A. 68, 2954–2957 (1971).

  3. 3

    Lyamichev, V., Brow, M. A. D. & Dahlberg, J. E. Science 260, 778–783 (1993).

  4. 4

    Harrington, J. J. & Lieber, M. R. EMBO J. 13, 1235–1246 (1994).

  5. 5

    Gutman, P. D. & Minton, K. W. Nucleic Acids Res. 21, 4406–4407 (1993).

  6. 6

    Sayers, J. R. & Eckstein, F. J. biol. Chem. 265, 18311–18317 (1990).

  7. 7

    Ceska, T. A., Sayers, J. R., Eckstein, F. & Suck, D. J. molec. Biol. 233, 179–182 (1993).

  8. 8

    Sayers, J. R. & Eckstein, F. Nucleic Acids Res. 19, 4127–4132 (1991).

  9. 9

    Kim, Y. et al. Nature 376, 612–616 (1995).

  10. 10

    Sayers, J. R. J. theor. Biol. 170, 415–421 (1994).

  11. 11

    Skinner, M. M. et al. Proc. natn. Acad. Sci. U.S.A. 91, 2071–2075 (1994).

  12. 12

    Beese, L. S. & Steitz, T. A. EMBO J. 10, 25–33 (1991).

  13. 13

    Lima, C. D., Wang, J. C. & Mondragon, A. Nature 367, 138–146 (1994).

  14. 14

    Berger, J. M., Gamblin, S. J., Harrison, S. C. & Wang, J. C. Nature 379, 225–233 (1996).

  15. 15

    Wigley, D. B., Davies, G. J., Dodson, E. J., Maxwell, A. & Dodson, G. Nature 351, 624–629 (1991).

  16. 16

    Robins, P., Pappin, D. J. C., Wood, R. D. & Lindahl, T. J. biol. Chem. 269, 28535–28538 (1994).

  17. 17

    Murante, R. S., Rust, L. & Bambara, R. A. J. biol. Chem. 270, 30377–30383 (1995).

  18. 18

    Sayers, J. R., Krekel, C. & Eckstein, F. BioTechniques 13, 592–596 (1992).

  19. 19

    Otwinowski, Z. DENZO (Yale University, New Haven, CT, 1993).

  20. 20

    Furey, W. & Swaminathan, S. Am. Crystallographic Ass. Meet. Abstr. 18, 73 (1990).

  21. 21

    Vellieux, F. M. D. A., Hunt, J. F., Roy, S. & Read, R. J. J. appl. Crystallogr. 28, 347–351 (1995).

  22. 22

    Jones, T. A., Zou, J.-Y., Cowan, S. W. & Kjeldgaard, M. Acta crystallogr. A47, 110–119 (1991).

  23. 23

    Collaborative Computational Project Number 4 Acta crystallogr. D50, 760–763 (1993).

  24. 24

    Brunger, A. T. X-PLOR, Version 3.1 (Yale University, New Haven, CT, 1993).

  25. 25

    Nicholls, A., Sharp, K. A. & Honig, B. Proteins Struct. Funct. Genet. 11, 282–296 (1991).

  26. 26

    Carson, M. J. molec. Graphics 5, 103–106 (1987).

Download references

Author information

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Ceska, T., Sayers, J., Stier, G. et al. A helical arch allowing single-stranded DNA to thread through T5 5'-exonuclease. Nature 382, 90–93 (1996). https://doi.org/10.1038/382090a0

Download citation

Further reading

Comments

By submitting a comment you agree to abide by our Terms and Community Guidelines. If you find something abusive or that does not comply with our terms or guidelines please flag it as inappropriate.