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A synthetic redox biofilm made from metalloprotein–prion domain chimera nanowires

Nature Chemistry volume 9, pages 157–163 (2017)Cite this article

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Abstract

Engineering bioelectronic components and set-ups that mimic natural systems is extremely challenging. Here we report the design of a protein-only redox film inspired by the architecture of bacterial electroactive biofilms. The nanowire scaffold is formed using a chimeric protein that results from the attachment of a prion domain to a rubredoxin (Rd) that acts as an electron carrier. The prion domain self-assembles into stable fibres and provides a suitable arrangement of redox metal centres in Rd to permit electron transport. This results in highly organized films, able to transport electrons over several micrometres through a network of bionanowires. We demonstrate that our bionanowires can be used as electron-transfer mediators to build a bioelectrode for the electrocatalytic oxygen reduction by laccase. This approach opens opportunities for the engineering of protein-only electron mediators (with tunable redox potentials and optimized interactions with enzymes) and applications in the field of protein-only bioelectrodes.

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Figure 1: Structural features of Rd-HET nanowires and redox film.
Figure 2: Electrochemistry of Rd-HET redox film.
Figure 3: EIS on Rd-HET redox film.
Figure 4: Rd-HET as a redox mediator for the bioelectrocatalytic reduction of oxygen by laccase.

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

    Article  CAS  PubMed  Google Scholar 

  2. Qian, F. & Li, Y. Biomaterials: a natural source of nanowires. Nat. Nanotechnol. 6, 538–539 (2011).

    Article  CAS  PubMed  Google Scholar 

  3. Pfeffer, C. et al. Filamentous bacteria transport electrons over centimetre distances. Nature 491, 218–221 (2012).

    Article  CAS  PubMed  Google Scholar 

  4. Reardon, P. N. & Mueller, K. T. Structure of the type IVa major pilin from the electrically conductive bacterial nanowires of Geobacter sulfurreducens. J. Biol. Chem. 288, 29260–29266 (2013).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. 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).

    Article  CAS  Google Scholar 

  6. Malvankar, N. S., Tuominen, M. T. & Lovley, D. R. Comment on ‘On electrical conductivity of microbial nanowires and biofilms’. Energy Environ. Sci. 5, 6247–6249 (2012).

    Article  CAS  Google Scholar 

  7. Bonanni, P. S., Massazza, D. & Busalmen, J. P. Stepping stones in the electron transport from cells to electrodes in Geobacter sulfurreducens biofilms. Phys. Chem. Chem. Phys. 15, 10300–10306 (2013).

    Article  CAS  PubMed  Google Scholar 

  8. Strycharz-Glaven, S. M., Snider, R. M., Guiseppi-Elie, A. & Tender, L. M. On the electrical conductivity of microbial nanowires and biofilms. Energy Environ. Sci. 4, 4366–4379 (2011).

    Article  CAS  Google Scholar 

  9. Knowles, T. P. J. & Buehler, M. J. Nanomechanics of functional and pathological amyloid materials. Nat. Nanotechnol. 6, 469–479 (2011).

    Article  CAS  PubMed  Google Scholar 

  10. Mankar, S., Anoop, A., Sen, S. & Maji, S. K. Nanomaterials: amyloids reflect their brighter side. Nano Rev. 2, 6032–6043 (2011).

    Article  Google Scholar 

  11. Gras, S. L. in Advances in Chemical Engineering Vol. 35 (ed. Rudy, J. K.) 161–209 (Academic, 2009).

    Google Scholar 

  12. Scheibel, T. et al. Conducting nanowires built by controlled self-assembly of amyloid fibers and selective metal deposition. Proc. Natl Acad. Sci. USA 100, 4527–4532 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Kholkin, A., Amdursky, N., Bdikin, I., Gazit, E. & Rosenman, G. Strong piezoelectricity in bioinspired peptide nanotubes. ACS Nano 4, 610–614 (2010).

    Article  CAS  PubMed  Google Scholar 

  14. Amit, M. et al. Hybrid proton and electron transport in peptide fibrils. Adv. Funct. Mater. 24, 5873–5880 (2014).

    Article  CAS  Google Scholar 

  15. Berger, O. et al. Light-emitting self-assembled peptide nucleic acids exhibit both stacking interactions and Watson–Crick base pairing. Nat. Nanotechnol. 10, 353–360 (2015).

    Article  CAS  PubMed  Google Scholar 

  16. del Mercato, L. L. et al. Charge transport and intrinsic fluorescence in amyloid-like fibrils. Proc. Natl Acad. Sci. USA 104, 18019–18024 (2007).

    Article  PubMed  PubMed Central  Google Scholar 

  17. Amit, M., Cheng, G., Hamley, I. W. & Ashkenasy, N. Conductance of amyloid-based peptide filaments: structure & function relations. Soft Matter 8, 8690–8696 (2012).

    Article  CAS  Google Scholar 

  18. Creasey, R. C. G., Shingaya, Y. & Nakayama, T. Improved electrical conductance through self-assembly of bioinspired peptides into nanoscale fibers. Mater. Chem. Phys. 158, 52–59 (2015).

    Article  CAS  Google Scholar 

  19. Domigan, L. J., Healy, J. P., Meade, S. J., Blaikie, R. J. & Gerrard, J. A. Controlling the dimensions of amyloid fibrils: toward homogenous components for bionanotechnology. Biopolymers 97, 123–133 (2012).

    Article  CAS  PubMed  Google Scholar 

  20. Herland, A. et al. Electroactive luminescent self-assembled bio-organic nanowires: integration of semiconducting oligoelectrolytes within amyloidogenic proteins. Adv. Mater. 17, 1466–1471 (2005).

    Article  CAS  Google Scholar 

  21. Rizzo, A., Solin, N., Lindgren, L. J., Andersson, M. R. & Inganäs, O. White light with phosphorescent protein fibrils in OLEDs. Nano Lett. 10, 2225–2230 (2010).

    Article  CAS  PubMed  Google Scholar 

  22. Bolisetty, S., Adamcik, J., Heier, J. & Mezzenga, R. Amyloid directed synthesis of titanium dioxide nanowires and their applications in hybrid photovoltaic devices. Adv. Funct. Mater. 22, 3424–3428 (2012).

    Article  CAS  Google Scholar 

  23. Li, C., Adamcik, J. & Mezzenga, R. Biodegradable nanocomposites of amyloid fibrils and graphene with shape-memory and enzyme-sensing properties. Nat. Nanotechnol. 7, 421–427 (2012).

    Article  CAS  PubMed  Google Scholar 

  24. Baldwin, A. J. et al. Cytochrome display on amyloid fibrils. J. Am. Chem. Soc. 128, 2162–2163 (2006).

    Article  CAS  PubMed  Google Scholar 

  25. Saupe, S. J. The [Het-s] prion of Podospora anserina and its role in heterokaryon incompatibility. Semin. Cell Dev. Biol. 22, 460–468 (2011).

    Article  CAS  PubMed  Google Scholar 

  26. Balguerie, A. et al. Domain organization and structure–function relationship of the HET-s prion protein of Podospora anserina. EMBO J. 22, 2071–2081 (2003).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  27. Wasmer, C. et al. Amyloid fibrils of the HET-s(218–289) prion form a β solenoid with a triangular hydrophobic core. Science 319, 1523–1526 (2008).

    Article  CAS  PubMed  Google Scholar 

  28. Meyer, J. Iron–sulfur protein folds, iron–sulfur chemistry, and evolution. J. Biol. Inorg. Chem. 13, 157–170 (2008).

    Article  CAS  PubMed  Google Scholar 

  29. Solomon, E. I., Sundaram, U. M. & Machonkin, T. E. Multicopper oxidases and oxygenases. Chem. Rev. 96, 2563–2606 (1996).

    Article  CAS  PubMed  Google Scholar 

  30. Van Melckebeke, H. et al. Atomic-resolution three-dimensional structure of HET-s(218–289) amyloid fibrils by solid-state NMR spectroscopy. J. Am. Chem. Soc. 132, 13765–13775 (2010).

    Article  CAS  PubMed  Google Scholar 

  31. Bonisch, H., Schmidt, C. L., Bianco, P. & Ladenstein, R. Ultrahigh-resolution study on Pyrococcus abyssi rubredoxin. I. 0.69 A X-ray structure of mutant W4L/R5S. Acta Cryst. D 61, 990–1004 (2005).

    Article  CAS  Google Scholar 

  32. Page, C. C., Moser, C. C., Chen, X. & Dutton, P. L. Natural engineering principles of electron tunnelling in biological oxidation-reduction. Nature 402, 47–52 (1999).

    Article  CAS  PubMed  Google Scholar 

  33. Winkler, J. R. & Gray, H. B. Long-range electron tunneling. J. Am. Chem. Soc. 136, 2930–2939 (2014).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Sabaté, R. et al. Prion and non-prion amyloids of the HET-s prion forming domain. J. Mol. Biol. 370, 768–783 (2007).

    Article  CAS  PubMed  Google Scholar 

  35. Doussineau, T. et al. Mass determination of entire amyloid fibrils by using mass spectrometry. Angew. Chem. Int. Ed. 55, 2340–2344 (2016).

    Article  CAS  Google Scholar 

  36. Siemer, A. et al. 13C, 15N resonance assignment of parts of the HET-s prion protein in its amyloid form. J. Biomol. NMR 34, 75–87 (2006).

    Article  CAS  PubMed  Google Scholar 

  37. Mizuno, N., Baxa, U. & Steven, A. C. Structural dependence of HET-s amyloid fibril infectivity assessed by cryoelectron microscopy. Proc. Natl Acad. Sci. USA 108, 3252–3257 (2011).

    Article  PubMed  PubMed Central  Google Scholar 

  38. Mao, F., Mano, N. & Heller, A. Long tethers binding redox centers to polymer backbones enhance electron transport in enzyme ‘wiring’ hydrogels. J. Am. Chem. Soc. 125, 4951–4957 (2003).

    Article  CAS  PubMed  Google Scholar 

  39. Bard, A. J. & Faulkner, L. R. in Electrochemical Methods: Fundamentals and Applications 2nd edn (eds Allen, J. & Bard, L. R. F.) 580 (John Wiley & Sons, 2001).

  40. Laviron, E. A multilayer model for the study of space distributed redox modified electrodes. Part I. Description and discussion of the model. J. Electroanal. Chem. Interfacial Electrochem. 112, 1–9 (1980).

    Article  CAS  Google Scholar 

  41. Blauch, D. N. & Saveant, J. M. Dynamics of electron hopping in assemblies of redox centers. Percolation and diffusion. J. Am. Chem. Soc. 114, 3323–3332 (1992).

    Article  CAS  Google Scholar 

  42. Andrieux, C. P. & Savéant, J. M. Electron transfer through redox polymer films. J. Electroanal. Chem. Interfacial Electrochem. 111, 377–381 (1980).

    Article  CAS  Google Scholar 

  43. Barsoukov, E. & Macdonald, J. R. Impedance Spectroscopy: Theory, Experiment, and Applications 2nd edn (John Wiley & Sons, 2005).

    Book  Google Scholar 

  44. Gabrielli, C., Haas, O. & Takenouti, H. Impedance analysis of electrodes modified with a reversible redox polymer film. J. Appl. Electrochem. 17, 82–90 (1987).

    Article  CAS  Google Scholar 

  45. Musiani, M. M. Characterization of electroactive polymer layers by electrochemical impedance spectroscopy (EIS). Electrochim. Acta 35, 1665–1670 (1990).

    Article  CAS  Google Scholar 

  46. Ho, C., Raistrick, I. D. & Huggins, R. A. Application of A-C techniques to the study of lithium diffusion in tungsten trioxide thin films. J. Electrochem. Soc. 127, 343–350 (1980).

    Article  CAS  Google Scholar 

  47. Bisquert, J., Garcia-Belmonte, G., Bueno, P., Longo, E. & Bulhões, L. O. S. Impedance of constant phase element (CPE)-blocked diffusion in film electrodes. J. Electroanal. Chem. 452, 229–234 (1998).

    Article  CAS  Google Scholar 

  48. Criado, C., Galán-Montenegro, P., Velásquez, P. & Ramos-Barrado, J. R. Diffusion with general boundary conditions in electrochemical systems. J. Electroanal. Chem. 488, 59–63 (2000).

    Article  CAS  Google Scholar 

  49. Ordinario, D. D. et al. Bulk protonic conductivity in a cephalopod structural protein. Nat. Chem. 6, 596–602 (2014).

    Article  CAS  PubMed  Google Scholar 

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

    Article  PubMed  Google Scholar 

  51. Rengaraj, S., Kavanagh, P. & Leech, D. A comparison of redox polymer and enzyme co-immobilization on carbon electrodes to provide membrane-less glucose/O2 enzymatic fuel cells with improved power output and stability. Biosens. Bioelectron. 30, 294–299 (2011).

    Article  CAS  PubMed  Google Scholar 

  52. Rubenwolf, S. et al. Carbon electrodes for direct electron transfer type laccase cathodes investigated by current density–cathode potential behavior. Biosens. Bioelectron. 26, 841–845 (2010).

    Article  CAS  PubMed  Google Scholar 

  53. Feifel, S. C., Kapp, A. & Lisdat, F. Electroactive nanobiomolecular architectures of laccase and cytochrome c on electrodes: applying silica nanoparticles as artificial matrix. Langmuir 30, 5363–5367 (2014).

    Article  CAS  PubMed  Google Scholar 

  54. Feifel, S. C., Kapp, A., Ludwig, R. & Lisdat, F. Nanobiomolecular multiprotein clusters on electrodes for the formation of a switchable cascadic reaction scheme. Angew. Chem. Int. Ed. 53, 5676–5679 (2014).

    Article  CAS  Google Scholar 

  55. Winkler, J. R., Gray, H. B., Prytkova, T. R., Kurnikov, I. V. & Beratan, D. N. in Bioelectronics: From Theory to Applications (eds Willner, I. & Katz, E.) 15–33 (Wiley-VCH, 2005).

    Book  Google Scholar 

  56. Piontek, K., Antorini, M. & Choinowski, T. Crystal structure of a laccase from the fungus Trametes versicolor at 1.90-Å resolution containing a full complement of coppers. J. Biol. Chem. 277, 37663–37669 (2002).

    Article  CAS  PubMed  Google Scholar 

  57. Le Goff, A., Holzinger, M. & Cosnier, S. Enzymatic biosensors based on SWCNT-conducting polymer electrodes. Analyst 136, 1279–1287 (2011).

    Article  CAS  PubMed  Google Scholar 

  58. Holzinger, M., Le Goff, A. & Cosnier, S. Carbon nanotube/enzyme biofuel cells. Electrochim. Acta 82, 179–190 (2012).

    Article  CAS  Google Scholar 

  59. Daniels, J. S. & Pourmand, N. Label-free impedance biosensors: opportunities and challenges. Electroanalysis 19, 1239–1257 (2007).

    Article  CAS  PubMed  PubMed Central  Google Scholar 

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Acknowledgements

We thank D. Fenel and G. Schoehn from the IBS/UVHCI platform of the Partnership for Structural Biology in Grenoble (PSB/IBS) for the electron microscopy. We thank S. Saupe for the gift of pET24a(+)-HET-s(218-289)-His6 and S. Crouzy for his help in building the structural model. AFM measurements were performed on the Commissariat à l'énergie atomique (CEA) Minatec Nanocharacterization Platform (PFNC). The present work was also partially supported by the Labex ARCANE (ANR-11-LABX-0003-01). L.A. and A.R are indebted to CEA for the funding of their PhD fellowships. V.F. and P.R. thank N. Mermilliod and E. Molva, respectively heads of the Transverse Energy and the Nanoscience programs of the CEA, for their scientific and financial support. We thank T. Martin for the careful reading of the manuscript.

Author information

Author notes

  1. Saravanan Rengaraj & Anthony L. B. Maçon

    Present address: Present addresses: Department of Chemical Engineering, University of Bath, Bath BA2 7AY, UK (S.R.); Nagoya Institute of Technology, Frontier Research Institute, Nagoya, Aichi 466-8555, Japan (A.L.B.M.),

  2. Lucie Altamura, Christophe Horvath and Saravanan Rengaraj: These authors contributed equally to this work

Authors and Affiliations

  1. Université Grenoble Alpes, BIG-LCBM, Grenoble, F-38000, France

    Lucie Altamura, Christophe Horvath, Saravanan Rengaraj, Anaëlle Rongier, Anthony L. B. Maçon, Marc Fontecave, Nicolas Duraffourg & Vincent Forge

  2. CNRS, BIG-LCBM, Grenoble, F-38000, France

    Lucie Altamura, Christophe Horvath, Saravanan Rengaraj, Anaëlle Rongier, Anthony L. B. Maçon, Marc Fontecave, Nicolas Duraffourg & Vincent Forge

  3. CEA, BIG-LCBM, Grenoble, F-38000, France

    Lucie Altamura, Christophe Horvath, Saravanan Rengaraj, Anaëlle Rongier, Anthony L. B. Maçon, Marc Fontecave, Nicolas Duraffourg & Vincent Forge

  4. Université Grenoble Alpes, DCM UMR 5250, Grenoble, F-38000, France

    Saravanan Rengaraj, Kamal Elouarzaki, Chantal Gondran, Alan Le Goff & Michael Holzinger

  5. CNRS, DCM UMR 5250, Grenoble, F-38000, France

    Saravanan Rengaraj, Kamal Elouarzaki, Chantal Gondran, Alan Le Goff & Michael Holzinger

  6. Université Grenoble Alpes, INAC-SPrAM, Grenoble, F-38000, France

    Anaëlle Rongier & Patrice Rannou

  7. CNRS, INAC-SPrAM, Grenoble, F-38000, France

    Anaëlle Rongier & Patrice Rannou

  8. CEA, INAC-SPrAM, Grenoble, F-38000, France

    Anaëlle Rongier & Patrice Rannou

  9. ERRMECe, I-MAT FD4122, Université de Cergy-Pontoise, 2 avenue Adolphe Chauvin, 95302 Cergy-Pontoise Cedex, France,

    Charlotte Vendrely

  10. LMGP, CNRS UMR 5628, 3 parvis Louis Néel, Grenoble, 38016, France

    Charlotte Vendrely

  11. Institut Néel, CNRS-UJF-INP, Grenoble Cedex 09, 38042, France

    Vincent Bouchiat

  12. Laboratoire de Chimie des Processus Biologiques, UMR 8229 CNRS, Université Pierre et Marie Curie – Paris 6, Collège de France. 11 Place Marcelin Berthelot, Paris Cedex 05, 75231, France

    Marc Fontecave

  13. CEA, LETI, MINATEC Campus, Grenoble, F-38054, France.

    Denis Mariolle

Authors
  1. Lucie Altamura
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Contributions

C.H., N.D., M.F. and V.F. realized the design of the protein nanowire. C.H., L.A. and N.D. developed the protocol for the chimeric protein production in bacteria and purification. C.H., L.A., C.V., D.M., V.B., N.D. and V.F. performed the biophysical characterization of the bionanowires. S.R., L.A., K.E., C.G., A.L.B.M., A.L.-G., M.H. and N.D. carried out the electrochemical characterization of the bionanowires and L.A., A.R. and P.R. performed the electrical characterizations of the bionanowires. S.R., K.E., A.L.-G. and M.H. designed the electrode functionalized with nanowires and laccase and realized the experiments. M.F., V.B., M.H. and V.F. were responsible for the project management. L.A., S.R., A.L.-G. and V.F. prepared the manuscript. All the authors discussed the results and commented on the manuscript.

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Correspondence to Michael Holzinger or Vincent Forge.

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Altamura, L., Horvath, C., Rengaraj, S. et al. A synthetic redox biofilm made from metalloprotein–prion domain chimera nanowires. Nature Chem 9, 157–163 (2017). https://doi.org/10.1038/nchem.2616

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