Letter | Published:

Design of a hyperstable 60-subunit protein icosahedron

Nature volume 535, pages 136139 (07 July 2016) | Download Citation

  • A Corrigendum to this article was published on 19 October 2016

This article has been updated

Abstract

The icosahedron is the largest of the Platonic solids, and icosahedral protein structures are widely used in biological systems for packaging and transport1,2. There has been considerable interest in repurposing such structures3,4,5 for applications ranging from targeted delivery to multivalent immunogen presentation. The ability to design proteins that self-assemble into precisely specified, highly ordered icosahedral structures would open the door to a new generation of protein containers with properties custom-tailored to specific applications. Here we describe the computational design of a 25-nanometre icosahedral nanocage that self-assembles from trimeric protein building blocks. The designed protein was produced in Escherichia coli, and found by electron microscopy to assemble into a homogenous population of icosahedral particles nearly identical to the design model. The particles are stable in 6.7 molar guanidine hydrochloride at up to 80 degrees Celsius, and undergo extremely abrupt, but reversible, disassembly between 2 molar and 2.25 molar guanidinium thiocyanate. The icosahedron is robust to genetic fusions: one or two copies of green fluorescent protein (GFP) can be fused to each of the 60 subunits to create highly fluorescent ‘standard candles’ for use in light microscopy, and a designed protein pentamer can be placed in the centre of each of the 20 pentameric faces to modulate the size of the entrance/exit channels of the cage. Such robust and customizable nanocages should have considerable utility in targeted drug delivery6, vaccine design7 and synthetic biology8.

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Change history

  • 06 July 2016

    An addition was made to the Acknowledgements section.

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Acknowledgements

This work was supported by the Howard Hughes Medical Institute (D.B. and T.G.), the JRC visitor programme (S.G.), the National Science Foundation CHE-1332907 (D.B.), a UW/Hutch CCSG Pilot Award NCI 5 P30 CA015704-41 (D.B. and N.P.K.), Takeda Pharmaceutical Company (N.P.K.), the Bill and Melinda Gates Foundation OPP1120319 (D.B. and N.P.K.), the National Institutes of Health (NIH) P41 GM103533 (T.N.D.), the Defense Advanced Research Projects Agency (D.B. and N.P.K., grant no. W911NF-14-1-0162) and the Air Force Office of Scientific Research (AFOSR) AFOSR FA950-12-10112 (D.B.). Y.H. was supported in part by a NIH Molecular Biology Training Grant (T32GM008268). U.N. was supported in part by a PHS National Research Service Award (T32GM007270) from NIGMS. J.B.B. was supported in part by an NSF Graduate Research Fellowship (DGE-0718124). We thank the Janelia Research Campus Cryo-EM Facility and J. de la Cruz for their assistance with the Titan Krios.

Author information

Author notes

    • Yang Hsia
    •  & Jacob B. Bale

    These authors contributed equally to this work.

Affiliations

  1. Department of Biochemistry, University of Washington, Seattle, Washington 98195, USA

    • Yang Hsia
    • , Jacob B. Bale
    • , Shane Gonen
    • , William Sheffler
    • , Kimberly K. Fong
    • , Una Nattermann
    • , Chunfu Xu
    • , Po-Ssu Huang
    • , Rashmi Ravichandran
    • , Sue Yi
    • , Trisha N. Davis
    • , Neil P. King
    •  & David Baker
  2. Institute for Protein Design, University of Washington, Seattle, Washington 98195, USA

    • Yang Hsia
    • , Jacob B. Bale
    • , Shane Gonen
    • , William Sheffler
    • , Una Nattermann
    • , Chunfu Xu
    • , Po-Ssu Huang
    • , Rashmi Ravichandran
    • , Sue Yi
    • , Neil P. King
    •  & David Baker
  3. Graduate Program in Biological Physics, Structure and Design, University of Washington, Seattle, Washington 98195, USA

    • Yang Hsia
    • , Shane Gonen
    •  & Una Nattermann
  4. Graduate Program in Molecular and Cellular Biology, University of Washington, Seattle, Washington 98195, USA

    • Jacob B. Bale
  5. Janelia Research Campus, Howard Hughes Medical Institute, Ashburn, Virginia 20147, USA

    • Shane Gonen
    • , Dan Shi
    •  & Tamir Gonen
  6. Howard Hughes Medical Institute, University of Washington, Seattle, Washington 98195, USA

    • David Baker

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Contributions

J.B.B., N.P.K., and W.S. developed the computational design methodology. Y.H. and J.B.B. performed the design of the icosahedra. Y.H. performed all other unlisted experiments. S.G. and D.S. performed the cryo-EM experiments. K.K.F. performed the fluorescence microscopy experiments. U.N. performed the negative-stain electron microscopy experiments. C.X. provided the pentamer sequence for I3-01(HB). P.-S.H. created the computational methodology to model fusions to I3-01. R.R. produced I3-01(HB) proteins. S.Y. produced T33-21 sfGFP fusion proteins.

Competing interests

The authors declare no competing financial interests.

Corresponding author

Correspondence to David Baker.

Extended data

Supplementary information

Zip files

  1. 1.

    Supplementary Information

    This zipped file contains the protein sequences, the design structure PDB file and an example script.

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DOI

https://doi.org/10.1038/nature18010

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