Visualization of charge propagation along individual pili proteins using ambient electrostatic force microscopy

Abstract

The nanoscale imaging of charge flow in proteins is crucial to understanding several life processes, including respiration, metabolism and photosynthesis1,2,3. However, existing imaging methods are only effective under non-physiological conditions or are limited to photosynthetic proteins1. Here, we show that electrostatic force microscopy can be used to directly visualize charge propagation along pili of Geobacter sulfurreducens with nanometre resolution and under ambient conditions. Charges injected at a single point into individual, untreated pili, which are still attached to cells, propagated over the entire filament. The mobile charge density in the pili, as well as the temperature and pH dependence of the charge density, were similar to those of carbon nanotubes4 and other organic conductors5,6,7. These findings, coupled with a lack of charge propagation in mutated pili that were missing key aromatic amino acids8, suggest that the pili of G. sulfurreducens function as molecular wires with transport via delocalized charges, rather than the hopping mechanism that is typical of biological electron transport2,3,9.

Access options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

Figure 1: Strategy for direct visualization of charge propagation along native bacterial proteins with ambient EFM.
Figure 2: EFM imaging demonstrates charge propagation along pili filaments.
Figure 3: Visualization of charge propagation along pili filaments.
Figure 4: Quantitative measurements of charge propagation in pili filaments with EFM.

References

  1. 1

    Plumere, N. Single molecules: a protein in the spotlight. Nature Nanotech. 7, 616–617 (2012).

    CAS  Article  Google Scholar 

  2. 2

    Edwards, P. P., Gray, H. B., Lodge, M. T. J. & Williams, R. J. P. Electron transfer and electronic conduction through an intervening medium. Angew. Chem. Int. Ed. 47, 6758–6765 (2008).

    CAS  Article  Google Scholar 

  3. 3

    Waleed Shinwari, M., Jamal Deen, M., Starikov, E. B. & Cuniberti, G. Electrical conductance in biological molecules. Adv. Funct. Mater. 20, 1865–1883 (2010).

    CAS  Article  Google Scholar 

  4. 4

    Melin, T., Zdrojek, M. & Brunel, D. in Scanning Probe Microscopy in Nanoscience and Nanotechnology (ed. Bhushan, B.) 89–128 (Springer, 2010).

  5. 5

    Dautel, O. J. et al. Electroactive nanorods and nanorings designed by supramolecular association of π-conjugated oligomers. Chem. Eur. J. 14, 4201–4213 (2008).

    CAS  Article  Google Scholar 

  6. 6

    Wang, S. et al. Hopping transport and the Hall effect near the insulator–metal transition in electrochemically gated poly(3-hexylthiophene) transistors. Nature Commun. 3, 1210 (2012).

    Article  Google Scholar 

  7. 7

    Heim, T., Lmimouni, K. & Vuillaume, D. Ambipolar charge injection and transport in a single pentacene monolayer island. Nano Lett. 4, 2145–2150 (2004).

    CAS  Article  Google Scholar 

  8. 8

    Vargas, M. et al. Aromatic amino acids required for pili conductivity and long-range extracellular electron transport in Geobacter sulfurreducens. mBio 4, e00105 (2013).

    Article  Google Scholar 

  9. 9

    Malvankar, N. S. & Lovley, D. R. Microbial nanowires for bioenergy applications. Curr. Opin. Biotechnol. 27, 88–95 (2014).

    CAS  Article  Google Scholar 

  10. 10

    Malvankar, N. S. & Lovley, D. R. Microbial nanowires: a new paradigm for biological electron transfer and bioelectronics. ChemSusChem 5, 1039–1046 (2012).

    CAS  Article  Google Scholar 

  11. 11

    Boesen, T. & Nielsen, L. P. Molecular dissection of bacterial nanowires. mBio 4, e00270 (2013).

    CAS  Article  Google Scholar 

  12. 12

    Wanger, G. et al. Electrically conductive bacterial nanowires in bisphosphonate-related osteonecrosis of the jaw biofilms. Oral Surg. Oral Med. Oral Pathol. Oral Radiol. 115, 71–78 (2013).

    Article  Google Scholar 

  13. 13

    Reguera, G. et al. Extracellular electron transfer via microbial nanowires. Nature 435, 1098–1101 (2005).

    CAS  Article  Google Scholar 

  14. 14

    Summers, Z. M. et al. Direct exchange of electrons within aggregates of an evolved syntrophic coculture of anaerobic bacteria. Science 330, 1413–1415 (2010).

    CAS  Article  Google Scholar 

  15. 15

    Leung, K. M. et al. Shewanella oneidensis MR-1 bacterial nanowires exhibit p-type, tunable electronic behavior. Nano Lett. 13, 2407–2411 (2013).

    CAS  Article  Google Scholar 

  16. 16

    Leang, C., Qian, X., Mester, T. & Lovley, D. R. Alignment of the c-type cytochrome OmcS along pili of Geobacter sulfurreducens. Appl. Environ. Microbiol. 76, 4080–4084 (2010).

    CAS  Article  Google Scholar 

  17. 17

    Malvankar, N. S., Tuominen, M. T. & Lovley, D. R. Lack of cytochrome involvement in long-range electron transport through conductive biofilms and nanowires of Geobacter sulfurreducens. Energy Environ. Sci. 5, 8651–8659 (2012).

    CAS  Article  Google Scholar 

  18. 18

    Veazey, J. P., Reguera, G. & Tessmer, S. H. Electronic properties of conductive pili of the metal-reducing bacterium Geobacter sulfurreducens probed by scanning tunneling microscopy. Phys. Rev. E 84, 060901 (2011).

    Article  Google Scholar 

  19. 19

    Malvankar, N. S. et al. Tunable metallic-like conductivity in microbial nanowire networks. Nature Nanotech. 6, 573–579 (2011).

    Article  Google Scholar 

  20. 20

    Dallas, P. et al. Characterization, magnetic and transport properties of polyaniline synthesized through interfacial polymerization. Polymer 48, 3162–3169 (2007).

    CAS  Article  Google Scholar 

  21. 21

    Chiang, J. C. & MacDiarmid, A. G. ‘ Polyaniline’: protonic acid doping of the emeraldine form to the metallic regime. Synth. Met. 13, 193–205 (1986).

    CAS  Article  Google Scholar 

  22. 22

    Dukovic, G. et al. Reversible surface oxidation and efficient luminescence quenching in semiconductor single-wall carbon nanotubes. J. Am. Chem. Soc. 126, 15269–15276 (2004).

    CAS  Article  Google Scholar 

  23. 23

    Mohn, F., Gross, L., Moll, N. & Meyer, G. Imaging the charge distribution within a single molecule. Nature Nanotech. 7, 227–231 (2012).

    CAS  Article  Google Scholar 

  24. 24

    Yalcin, S. E., Labastide, J. A., Sowle, D. L. & Barnes, M. D. Spectral properties of multiply charged semiconductor quantum dots. Nano Lett. 11, 4425–4430 (2011).

    CAS  Article  Google Scholar 

  25. 25

    Yalcin, S. E., Yang, B., Labastide, J. A. & Barnes, M. D. Electrostatic force microscopy and spectral studies of electron attachment to single quantum dots on indium tin oxide substrates. J. Phys. Chem. C 116, 15847–15853 (2012).

    CAS  Article  Google Scholar 

  26. 26

    Malvankar, N. S., Mester, T., Tuominen, M. T. & Lovley, D. R. Supercapacitors based on c-type cytochromes using conductive nanostructured networks of living bacteria. ChemPhysChem 13, 463–468 (2012).

    CAS  Article  Google Scholar 

  27. 27

    Medalsy, I. et al. Logic implementations using a single nanoparticle–protein hybrid. Nature Nanotech. 5, 451–457 (2010).

    CAS  Article  Google Scholar 

  28. 28

    Bube, R. H. Electrons in Solids: an Introductory Survey (Academic, 1992).

  29. 29

    Fumagalli, L. et al. Label-free identification of single dielectric nanoparticles and viruses with ultraweak polarization forces. Nature Mater. 11, 808–816 (2012).

    CAS  Article  Google Scholar 

  30. 30

    Schutz, C. N. & Warshel, A. What are the dielectric ‘constants' of proteins and how to validate electrostatic models? Prot. Struct. Funct. Bioinform. 44, 400–417 (2001).

    CAS  Article  Google Scholar 

Download references

Acknowledgements

This research was supported by the Office of Naval Research (grant nos N00014-13-1-0550 and N00014-12-1-0229), the Office of Science (BER), US Department of Energy (award no. DE-SC0006790) and the National Science Foundation Centre for Hierarchical Manufacturing (grant no. CMMI-1025020). The authors thank A. Parsegian for discussions, Asylum Research for technical support, M. Vargas and M. Sharma for help with cell culturing, D. Callaham for help with TEM and S. Thirunavukkarasu and T. Emrick for use of the W.M. Keck Nanostructures Facility (University of Massachusetts Amherst) for EFM studies. The Asylum Research MFP-3D equipment used for EFM was purchased via the Polymer-Based Materials for Harvesting Solar Energy and Energy Frontier Research Centre funded by the US Department of Energy, Office of Science, Office of Basic Energy Sciences (grant no. DE-SC0001087). N.S.M. holds a Career Award from the Scientific Interface from the Burroughs Wellcome Fund.

Author information

Affiliations

Authors

Contributions

N.S.M. and S.E.Y. conceived, designed and performed the experiments and analysed the data. M.T.T. and D.R.L. supervised the project. N.S.M. and D.R.L prepared the manuscript with critical comments from all authors.

Corresponding authors

Correspondence to Nikhil S. Malvankar or Derek R. Lovley.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Information

Supplementary Information (PDF 519 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Malvankar, N., Yalcin, S., Tuominen, M. et al. Visualization of charge propagation along individual pili proteins using ambient electrostatic force microscopy. Nature Nanotech 9, 1012–1017 (2014). https://doi.org/10.1038/nnano.2014.236

Download citation

Further reading

Search

Find nanotechnology articles, nanomaterial data and patents all in one place. Visit Nano by Nature Research