Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Letter
  • Published:

The bacteriophage φ29 portal motor can package DNA against a large internal force

Nature volume 413, pages 748–752 (2001)Cite this article

Abstract

As part of the viral infection cycle, viruses must package their newly replicated genomes for delivery to other host cells. Bacteriophage φ29 packages its 6.6-µm long, double-stranded DNA into a 42 × 54 nm capsid1 by means of a portal complex that hydrolyses ATP2. This process is remarkable because entropic, electrostatic and bending energies of the DNA must be overcome to package the DNA to near-crystalline density. Here we use optical tweezers to pull on single DNA molecules as they are packaged, thus demonstrating that the portal complex is a force-generating motor. This motor can work against loads of up to 57 pN on average, making it one of the strongest molecular motors reported to date. Movements of over 5 µm are observed, indicating high processivity. Pauses and slips also occur, particularly at higher forces. We establish the force–velocity relationship of the motor and find that the rate-limiting step of the motor's cycle is force dependent even at low loads. Notably, the packaging rate decreases as the prohead is filled, indicating that an internal force builds up to ∼50 pN owing to DNA confinement. Our data suggest that this force may be available for initiating the ejection of the DNA from the capsid during infection.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1: Set-up and initial results.
Figure 2: Measurements of packaging in no feedback mode.
Figure 3: Force–velocity (F-v) analysis.

Similar content being viewed by others

References

  1. Tao, Y. et al. Assembly of a tailed bacterial virus and its genome release studied in three dimensions. Cell 95, 431–437 (1998).

    Article  CAS  Google Scholar 

  2. Guo, P., Peterson, C. & Anderson, D. Prohead and DNA-gp3-dependent ATPase activity of the DNA packaging protein gp16 of bacteriophage φ29. J. Mol. Biol. 197, 229–236 (1987).

    Article  CAS  Google Scholar 

  3. Anderson, D. & Reilly, B. in Bacillus subtilis and Other Gram-Positive Bacteria: Biochemistry, Physiology, and Molecular Genetics (eds Sonenshein, A., Hoch, J. A. & Losick, R.) 859–867 (American Society for Microbiology, Washington DC, 1993).

    Google Scholar 

  4. Guo, P., Grimes, S. & Anderson, D. A defined system for in vitro packaging of DNA-gp3 of the Bacillus subtilis bacteriophage φ29. Proc. Natl Acad. Sci. USA 83, 3505–3509 (1986).

    Article  ADS  CAS  Google Scholar 

  5. Grimes, S. & Anderson, D. In vitro packaging of bacteriophage φ29 DNA restriction fragments and the role of the terminal protein gp3. J. Mol. Biol. 209, 91–100 (1989).

    Article  CAS  Google Scholar 

  6. Simpson, A. A. et al. Structure of the bacteriophage φ29 DNA packaging motor. Nature 408, 745–750 (2000).

    Article  ADS  CAS  Google Scholar 

  7. Ibarra, B. et al. Topology of the components of the DNA packaging machinery in the phage φ29 prohead. J. Mol. Biol. 298, 807–815 (2000).

    Article  CAS  Google Scholar 

  8. Smith, S. B., Cui, Y. & Bustamante, C. Overstretching B-DNA: the elastic response of individual double-stranded and single-stranded DNA molecules. Science 271, 795–799 (1996).

    Article  ADS  CAS  Google Scholar 

  9. Barlow, R. Statistics: a Guide to the Use of Statistical Methods in the Physical Sciences 15–16 (Wiley, Chichester, 1989).

    MATH  Google Scholar 

  10. Wang, M. D. et al. Force and velocity measured for single molecules of RNA polymerase. Science 282, 902–907 (1998).

    Article  ADS  CAS  Google Scholar 

  11. Berg, H. C. & Turner, L. Torque generated by the flagellar motor of Escherichia coli. Biophys. J. 65, 2201–2216 (1993).

    Article  ADS  CAS  Google Scholar 

  12. Kramers, H. Brownian motion in a field of force and the diffusion model of chemical reactions. Physica 7, 284–304 (1940).

    Article  ADS  MathSciNet  CAS  Google Scholar 

  13. Svoboda, K. & Block, S. M. Force and velocity measured for single kinesin molecules. Cell 77, 773–784 (1994).

    Article  CAS  Google Scholar 

  14. Coppin, C. M., Pierce, D. W., Hsu, L. & Vale, R. D. The load dependence of kinesin's mechanical cycle. Proc. Natl Acad. Sci. USA 94, 8539–8544 (1997).

    Article  ADS  CAS  Google Scholar 

  15. Finer, J. T., Simmons, R. M. & Spudich, J. A. Single myosin molecule mechanics: piconewton forces and nanometre steps. Nature 368, 113–119 (1994).

    Article  ADS  CAS  Google Scholar 

  16. Lehninger, A. L., Nelson, D. L. & Cox, M. M. Principles of Biochemistry 375 (Worth, New York, 1993).

    Google Scholar 

  17. Earnshaw, W. C. & Casjens, S. R. DNA packaging by the double-stranded DNA bacteriophages. Cell 21, 319–331 (1980).

    Article  CAS  Google Scholar 

  18. Rau, D. C., Lee, B. & Parsegian, V. A. Measurement of the repulsive force between polyelectrolyte molecules in ionic solution: hydration forces between parallel DNA double helices. Proc. Natl Acad. Sci. USA 81, 2621–2625 (1984).

    Article  ADS  CAS  Google Scholar 

  19. Riemer, S. C. & Bloomfield, V. A. Packaging of DNA in bacteriophage heads: some considerations on energetics. Biopolymers 17, 785–794 (1978).

    Article  CAS  Google Scholar 

  20. Odijk, T. Hexagonally packed DNA within bacteriophage T7 stabilized by curvature stress. Biophys. J. 75, 1223–1227 (1998).

    Article  ADS  CAS  Google Scholar 

  21. Garcia, L. R. & Molineux, I. J. Transcription-independent DNA translocation of bacteriophage T7 DNA into Escherichia coli. J. Bacteriol. 178, 6921–6929 (1996).

    Article  CAS  Google Scholar 

  22. Novick, S. L. & Baldeschwieler, J. D. Fluorescence measurement of the kinetics of DNA injection by bacteriophage lambda into liposomes. Biochemistry 27, 7919–7924 (1988).

    Article  CAS  Google Scholar 

  23. Grimes, S. & Anderson, D. The bacteriophage φ29 packaging proteins supercoil the DNA ends. J. Mol. Biol. 266, 901–914 (1997).

    Article  CAS  Google Scholar 

  24. Bjornsti, M. A., Reilly, B. E. & Anderson, D. L. Morphogenesis of bacteriophage φ29 of Bacillus subtilis: oriented and quantized in vitro packaging of DNA-gp3. J. Virol. 45, 383–396 (1983).

    CAS  PubMed  PubMed Central  Google Scholar 

  25. Baumann, C. G., Smith, S. B., Bloomfield, V. A. & Bustamante, C. Ionic effects on the elasticity of single DNA molecules. Proc. Natl Acad. Sci. USA 94, 6185–6190 (1997).

    Article  ADS  CAS  Google Scholar 

Download references

Acknowledgements

We thank W. M. Gelbart, P. Jardine, T. Odijk, V. Bloomfield, D. Frenkel, C. Varga, A. Mehta and M. Young for comments. This research was supported in part by grants from the NIH, DOE and NSF. D.E.S. and S.J.T. are supported by a grant from the Packard Foundation. S.J.T. is supported by the Netherlands Organization for Scientific Research (NWO).

Author information

Author notes

  1. Douglas E. Smith, Sander J. Tans and Carlos Bustamante: These authors contributed equally to this work

Authors and Affiliations

  1. Department of Physics, University of California, Berkeley, 94720, California, USA

    Douglas E. Smith, Sander J. Tans & Carlos Bustamante

  2. Department of Molecular and Cell Biology, University of California, Berkeley, 94720, California, USA

    Carlos Bustamante

  3. Howard Hughes Medical Institute, University of California, Berkeley, 94720, California, USA

    Steven B. Smith & Carlos Bustamante

  4. Physical Biosciences Division of Lawrence Berkeley Laboratory, University of California, Berkeley, 94720, California, USA

    Carlos Bustamante

  5. Departments of Microbiology and Oral Science, University of Minnesota, Minneapolis, 55455, Minnesota, USA

    Shelley Grimes & Dwight L. Anderson

Authors
  1. Douglas E. Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sander J. Tans
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Steven B. Smith
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Shelley Grimes
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Dwight L. Anderson
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Carlos Bustamante
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Carlos Bustamante.

Supplementary information

Rights and permissions

Reprints and permissions

About this article

Cite this article

Smith, D., Tans, S., Smith, S. et al. The bacteriophage φ29 portal motor can package DNA against a large internal force. Nature 413, 748–752 (2001). https://doi.org/10.1038/35099581

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/35099581

This article is cited by

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.

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing