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Mapping protein binding sites on the biomolecular corona of nanoparticles

Nature Nanotechnology volume 10, pages 472–479 (2015)Cite this article

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Abstract

Nanoparticles in a biological milieu are known to form a sufficiently long-lived and well-organized ‘corona’ of biomolecules to confer a biological identity to the particle. Because this nanoparticle–biomolecule complex interacts with cells and biological barriers, potentially engaging with different biological pathways, it is important to clarify the presentation of functional biomolecular motifs at its interface. Here, we demonstrate that by using antibody-labelled gold nanoparticles, differential centrifugal sedimentation and various imaging techniques it is possible to identify the spatial location of proteins, their functional motifs and their binding sites. We show that for transferrin-coated polystyrene nanoparticles only a minority of adsorbed proteins exhibit functional motifs and the spatial organization appears random, which is consistent, overall, with a stochastic and irreversible adsorption process. Our methods are applicable to a wide array of nanoparticles and can offer a microscopic molecular description of the biological identity of nanoparticles.

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Figure 1: Three-dimensional structure of transferrin (Tf) and the transferrin receptor (TfR)–transferrin complex.
Figure 2: Schematic representation of the use of DCS to measure immunogold labelling.
Figure 3: Immunogold labelling and epitope mapping of transferrin-coated 220 nm polystyrene nanoparticles.
Figure 4: Number and spatial organization of epitopes exposed on the surface of transferrin-coated 220 nm polystyrene nanoparticles.
Figure 5: Interaction of transferrin-coated 220 nm polystyrene nanoparticles with transferrin receptor.
Figure 6: Immunogold labelling of plasma-derived corona on 220 nm polystyrene nanoparticles.

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Acknowledgements

This work was supported by a Science Foundation Ireland (SFI) Principal Investigator Award (agreement no. 12/IA/1422). P.M.K. acknowledges the Irish Research Council for funding under the Embark scheme (RS/2011/106). The authors also acknowledge the small collaborative project NanoTransKinetics (NMP4-2010-EU-US-266737) funded by the European Union Seventh Framework Programme. This work was based on work supported by NanoSolutions (NMP-L6-2012-309329) and NAMDIATREAM (Nanotechnological Toolkit for Multi-modal Disease Diagnostics and Treatment Monitoring; NMP4-LA-2010-246479), both funded by the European Commission Seventh Framework Programme. The authors acknowledge support from Mass Spectrometry Resource and Conway Core Technologies, based at the Conway Institute of Biomolecular and Biomedical Research (University College Dublin), and the Advanced Microscopy Laboratory at the Centre for Research on Adaptive Nanostructures and Nanodevices (Trinity College Dublin) for access to advanced electron microscopy facilities.

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Authors and Affiliations

  1. Centre for BioNano Interactions, School of Chemistry and Chemical Biology, University College Dublin, Belfield, Dublin 4, Ireland

    Philip M. Kelly, Christoffer Åberg, Ester Polo, Ann O'Connell, Jennifer Cookman, Jonathan Fallon, Željka Krpetić & Kenneth A. Dawson

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  1. Philip M. Kelly
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  7. Željka Krpetić
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  8. Kenneth A. Dawson
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Contributions

P.M.K. designed and performed the experiments, collected, analysed and interpreted the data, and contributed to writing the manuscript. C.Å. constructed a theoretical framework to describe the experimental results and created the pairwise distributions, interpreted the data and contributed to writing the manuscript. E.P. contributed to the biological interaction experiments. A.O'C. performed the advanced electron microscopy analysis. J.C. contributed to TEM imaging. J.F. contributed to particle counting and image analysis. Z.K. contributed to the design of the experiments, oversaw the acquisition and interpretation of the microscopy images, and contributed to writing the manuscript. K.A.D. conceived and designed the experiments, interpreted the data and wrote the paper. All authors discussed the results and commented on the manuscript.

Corresponding authors

Correspondence to Željka Krpetić or Kenneth A. Dawson.

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The authors declare no competing financial interests.

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Kelly, P., Åberg, C., Polo, E. et al. Mapping protein binding sites on the biomolecular corona of nanoparticles. Nature Nanotech 10, 472–479 (2015). https://doi.org/10.1038/nnano.2015.47

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