Molecular Biotechnology

, Volume 57, Issue 9, pp 814–819 | Cite as

Xylan-Degrading Catalytic Flagellar Nanorods

  • Ágnes Klein
  • Veronika Szabó
  • Mátyás Kovács
  • Dániel Patkó
  • Balázs Tóth
  • Ferenc VondervisztEmail author


Flagellin, the main component of flagellar filaments, is a protein possessing polymerization ability. In this work, a novel fusion construct of xylanase A from B. subtilis and Salmonella flagellin was created which is applicable to build xylan-degrading catalytic nanorods of high stability. The FliC–XynA chimera when overexpressed in a flagellin deficient Salmonella host strain was secreted into the culture medium by the flagellum-specific export machinery allowing easy purification. Filamentous assemblies displaying high surface density of catalytic sites were produced by ammonium sulfate-induced polymerization. FliC–XynA nanorods were resistant to proteolytic degradation and preserved their enzymatic activity for a long period of time. Furnishing enzymes with self-assembling ability to build catalytic nanorods offers a promising alternative approach to enzyme immobilization onto nanostructured synthetic scaffolds.


Flagellin Xylanase A Polymerization Self-assembly Flagellar export Nanorod 





Xylanase A from Bacillus subtilis


Fusion construct of FliC and XynA with a C-terminal His6 tag


Sodium dodecyl sulfate–polyacrylamide gel electrophoresis


Phosphate buffered saline


Ammonium sulfate


Flagellin-enzyme fusion protein



We thank K. Namba and P. Závodszky for support and encouragement. We are grateful to the Japan Science and Technology Corporation for generous donation of equipment. This work was supported by the Hungarian Scientific Research Fund (OTKA) grants K104726 and NK108642.


  1. 1.
    Ge, J., Lu, D., Liu, Z., & Liu, Z. (2009). Recent advances in nanostructured biocatalysts. Biochemical Engineering Journal, 44, 53–59.CrossRefGoogle Scholar
  2. 2.
    Kim, J., Grate, J. W., & Wang, P. (2008). Nanobiocatalysis and its potential applications. Trends in Biotechnology, 26, 639–646.CrossRefGoogle Scholar
  3. 3.
    Govan, J., & Gun’ko, Y. K. (2014). Recent advances in the application of magnetic nanoparticles as a support for homogeneous catalysts. Nanomaterials, 4, 222–241.CrossRefGoogle Scholar
  4. 4.
    Misson, M., Zhang, H., & Jin, B. (2015). Nanobiocatalyst advancements and bioprocessing applications. Journal of Royal Society Interface, 12, 20140891.CrossRefGoogle Scholar
  5. 5.
    Talbert, J. N., & Goddard, J. M. (2012). Enzymes on material surfaces. Colloids and Surfaces B: Biointerfaces, 93, 8–19.CrossRefGoogle Scholar
  6. 6.
    Ansari, S. A., & Husain, Q. (2012). Potential applications of enzymes immobilized on/in nano materials: A review. Biotechnology Advances, 30, 512–523.CrossRefGoogle Scholar
  7. 7.
    Baxa, U., Speransky, V., Steven, A. C., & Wickner, R. B. (2002). Mechanism of inactivation on prion conversion of the Saccharomyces cerevisiae Ure2 protein. Proceedings of the National Academy of Sciences USA, 99, 5253–5260.CrossRefGoogle Scholar
  8. 8.
    Baldwin, A. J., Bader, R., Christodoulou, J., MacPhee, C. E., Dobson, C. M., & Barker, P. D. (2006). Cytochrome display on amyloid fibrils. Journal of the American Chemical Society, 128, 2162–2163.CrossRefGoogle Scholar
  9. 9.
    Szabó, V., Muskotál, A., Tóth, B., Mihovilovic, M. D., & Vonderviszt, F. (2011). Construction of a xylanase A variant capable of polymerization. PLoS One, 6, e25388.CrossRefGoogle Scholar
  10. 10.
    Vonderviszt, F., & Namba, K. (2008). Structure, function and assembly of flagellar axial proteins. In T. Scheibel (Ed.), Fibrous Proteins (pp. 58–76). Austin: Landes Biosciences.Google Scholar
  11. 11.
    Asakura, S. (1970). Polymerization of flagellin and polymorphism of flagella. Advances in Biophysics, 1, 99–155.Google Scholar
  12. 12.
    Kulkarni, N., Shendye, A., & Rao, M. (1999). Molecular and biotechnological aspects of xylanases. FEMS Microbiology Reviews, 23, 411–456.CrossRefGoogle Scholar
  13. 13.
    Muskotál, A., Seregélyes, C., Sebestyén, A., & Vonderviszt, F. (2010). Structural basis for stabilization of the hypervariable D3 domain of Salmonella flagellin upon filament formation. Journal of Molecular Biology, 403, 607–615.CrossRefGoogle Scholar
  14. 14.
    Klein, Á., Tóth, B., Jankovics, H., Muskotál, A., & Vonderviszt, F. (2011). A polymerizable GFP variant. Protein Engineering, Design & Selection, 25, 153–157.CrossRefGoogle Scholar
  15. 15.
    Vonderviszt, F., Kanto, S., Aizawa, S. I., & Namba, K. (1989). Terminal regions of flagellin are disordered in solution. Journal of Molecular Biology, 209, 127–133.CrossRefGoogle Scholar
  16. 16.
    Vonderviszt, F., Aizawa, S. I., & Namba, K. (1991). Role of the disordered terminal regions of flagellin in filament formation and stability. Journal of Molecular Biology, 221, 1461–1474.CrossRefGoogle Scholar
  17. 17.
    Minamino, T. (2014). Protein export through the bacterial type III export pathway. Biochimica et Biophysica Acta, 1843, 1642–1648.CrossRefGoogle Scholar
  18. 18.
    Gasteiger, E., Hoogland, C., Gattiker, A., Duvaud, S., Wilkins, M. R., et al. (2005). Protein identification and analysis tools on the ExPASy server. In J. M. Walker (Ed.), The proteomics protocols handbook (pp. 571–607). NJ: Humana Press.CrossRefGoogle Scholar
  19. 19.
    Bernfeld, P. (1955). Amylases α and β. Methods in Enzymology, 1, 149–158.CrossRefGoogle Scholar
  20. 20.
    Yonekura, K., Maki-Yonekura, S., & Namba, K. (2003). Complete atomic model of the bacterial flagellar filament by electron cryomicroscopy. Nature, 424, 643–650.CrossRefGoogle Scholar
  21. 21.
    Yonekura, K., Maki, S., Morgan, D. G., DeRosier, D. J., Vonderviszt, F., et al. (2000). The bacterial flagellar cap as the rotary promoter of flagellin self-assembly. Science, 290, 2148–2152.CrossRefGoogle Scholar
  22. 22.
    Végh, B. M., Gál, P., Dobó, J., Závodszky, P., & Vonderviszt, F. (2006). Localization of the flagellum-specific export signal in Salmonella flagellin. Biochemical and Biophysical Research Communications, 345, 93–98.CrossRefGoogle Scholar
  23. 23.
    Vonderviszt, F., Sajó, R., Dobó, J., & Závodszky, P. (2012). The use of a flagellar export signal for the secretion of recombinant proteins in Salmonella. Methods in Molecular Biology, 824, 131–143.CrossRefGoogle Scholar
  24. 24.
    Ohnishi, K., Ohto, Y., Aizawa, S. I., Macnab, R. M., & Iino, T. (1994). FlgD is a scaffolding protein needed for flagellar hook assembly in Salmonella typhimurium. Journal of Bacteriology, 176, 2272–2281.Google Scholar
  25. 25.
    Komoriya, K., Shibano, N., Higano, T., Azuma, N., Yamaguchi, S., & Aizawa, S. I. (1999). Flagellar proteins and type III-exported virulence factors are the predominant proteins secreted into the culture media of Salmonella typhimurium. Molecular Microbiology, 34, 767–779.CrossRefGoogle Scholar
  26. 26.
    Mimori-Kiyosue, Y., Vonderviszt, F., & Namba, K. (1997). Locations of terminal segments of flagellin in the filament structure and their roles in polymerization and polymorphism. Journal of Molecular Biology, 270, 222–237.CrossRefGoogle Scholar
  27. 27.
    Murakami, M. T., Arni, R. K., Vieira, D. S., Degreve, L., Ruller, R., & Ward, R. J. (2005). Correlation of temperature induced conformational change with optimum catalytic activity in the recombinant G/11 xylanase A from Bacillus subtilis strain 168 (1A1). FEBS Letters, 579, 6505–6510.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2015

Authors and Affiliations

  • Ágnes Klein
    • 1
  • Veronika Szabó
    • 1
  • Mátyás Kovács
    • 1
  • Dániel Patkó
    • 3
  • Balázs Tóth
    • 1
    • 2
  • Ferenc Vonderviszt
    • 1
    • 3
    Email author
  1. 1.Bio-Nanosystems Laboratory, Research Institute of Chemical and Process Engineering, Faculty of Information TechnologyUniversity of PannoniaVeszprémHungary
  2. 2.Agricultural Institute, Centre for Agricultural ResearchHungarian Academy of SciencesMartonvásárHungary
  3. 3.Institute for Technical Physics and Materials Science, Centre for Energy ResearchHungarian Academy of SciencesBudapestHungary

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