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A virus-based single-enzyme nanoreactor

Nature Nanotechnology volume 2, pages 635–639 (2007)Cite this article

Abstract

Most enzyme studies are carried out in bulk aqueous solution, at the so-called ensemble level, but more recently studies have appeared in which enzyme activity is measured at the level of a single molecule, revealing previously unseen properties1,2,3,4. To this end, enzymes have been chemically or physically anchored to a surface, which is often disadvantageous because it may lead to denaturation. In a natural environment, enzymes are present in a confined reaction space, which inspired us to develop a generic method to carry out single-enzyme experiments in the restricted spatial environment of a virus capsid. We report here the incorporation of individual horseradish peroxidase enzymes in the inner cavity of a virus, and describe single-molecule studies on their enzymatic behaviour. These show that the virus capsid is permeable for substrate and product and that this permeability can be altered by changing pH.

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Figure 1: Characterization of CCMV and the empty CCMV capsid.
Figure 2: Inclusion of HRP in the virus capsid.
Figure 3: Enzyme activity in the presence of single capsids.
Figure 4: Single-capsid experiments.

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Acknowledgements

This work was supported by the Council for Chemical Sciences of the Netherlands Organization for Scientific Research (NWO-CW) through a TOP grant to R.J.M.N. and Veni & Vidi Innovative Research Grants to J.J.L.M.C., and by the Royal Netherlands' Academy for Arts and Sciences (R.J.M.N.). The protein Dronpa was a generous gift from J. Hofkens, University of Leuven (Belgium).

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Authors and Affiliations

  1. Institute for Molecules and Materials, Radboud University Nijmegen, Toernooiveld 1, Nijmegen, 6525, ED, The Netherlands

    Marta Comellas-Aragonès, Hans Engelkamp, Victor I. Claessen, Alan E. Rowan, Peter C. M. Christianen, Jan C. Maan, Jeroen J. L. M. Cornelissen & Roeland J. M. Nolte

  2. Department of Biomedical Engineering, Laboratory of Macromolecular and Organic Chemistry and Soft Matter CryoTEM Research Unit, Eindhoven University of Technology, PO Box 513, Eindhoven, 5600, MB, The Netherlands

    Nico A. J. M. Sommerdijk

  3. Laboratory of Virology, Wageningen University, Binnenhaven 11, Wageningen, 6709, PD, The Netherlands

    Benedictus J. M. Verduin

Authors
  1. Marta Comellas-Aragonès
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Contributions

M.C., H.E., J.C. and R.N conceived and designed the experiments; M.C. carried out the capsid isolation and encapsulation experiments and M.C., V.C. and H.E. performed the fluorescence spectroscopy. B.V. contributed to the capsid isolation and encapsulation studies. H.E. analysed the fluorescence data. A.R., P.C., J.M. and R.N. facilitated the fluorescence microscope and N.S. provided the cryo-TEM data.M.C., H.E., J.C and R.N co-wrote the manuscript. All authors discussed the results and commented on the manuscript.

Corresponding authors

Correspondence to Hans Engelkamp or Jeroen J. L. M. Cornelissen.

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Comellas-Aragonès, M., Engelkamp, H., Claessen, V. et al. A virus-based single-enzyme nanoreactor. Nature Nanotech 2, 635–639 (2007). https://doi.org/10.1038/nnano.2007.299

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