Skip to main content

Modification of CCMV Nanocages for Enzyme Encapsulation

  • Protocol
  • First Online:

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1798))

Abstract

In cellular systems, compartmentalization plays an important role in the protection and regulation of enzymes. Controlled encapsulation of enzymes in nanocompartments is crucial in understanding biocatalytic processes in the cellular environment. We have recently described an enzymatic method to covalently attach enzymes, equipped with a small recognition peptide, to the interior of viral capsids. Viral capsids are especially interesting in this respect, as they form very well-defined nanoparticles with a uniform size and shape. Here, we describe the relevant experimental procedures to encapsulate a model enzyme into the interior of a viral capsid, purify the resulting viral capsids, and measure the catalytic activity of the encapsulated enzymes.

This is a preview of subscription content, log in via an institution.

Buying options

Protocol
USD   49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD   89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD   119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD   169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Learn about institutional subscriptions

Springer Nature is developing a new tool to find and evaluate Protocols. Learn more

References

  1. Schoonen L, van Hest JCM (2014) Functionalization of protein-based nanocages for drug delivery applications. Nanoscale 6:7124–7141

    Article  CAS  PubMed  Google Scholar 

  2. Caspar DLD, Klug A (1962) Physical principles in the construction of regular viruses. Cold Spring Harb Symp Quant Biol 27:1–24

    Article  CAS  PubMed  Google Scholar 

  3. Speir JA, Munshi S, Wang G et al (1995) Structures of the native and swollen forms of cowpea chlorotic mottle virus determined by X-ray crystallography and cryo-electron microscopy. Structure 3:63–78

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Schoonen L, Pille J, Borrmann A et al (2015) Sortase A-mediated N-terminal modification of cowpea chlorotic mottle virus for highly efficient cargo loading. Bioconjug Chem 26:2429–2434

    Article  CAS  PubMed  Google Scholar 

  5. Clancy KW, Melvin JA, McCafferty DG (2010) Sortase transpeptidases: insights into mechanism, substrate specificity, and inhibition. Biopolymers 94:385–396

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Schoonen L, Nolte RJM, van Hest JCM (2016) Highly efficient enzyme encapsulation in a protein nanocage: towards enzyme catalysis in a cellular nanocompartment mimic. Nanoscale 8:14467–14472

    Article  CAS  PubMed  Google Scholar 

  7. van Eldijk MB, Wang JC-Y, Minten IJ et al (2012) Designing two self-assembly mechanisms into one viral capsid protein. J Am Chem Soc 134:18506–18509

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  8. Gill SC, von Hippel PH (1989) Calculation of protein extinction coefficients from amino acid sequence data. Anal Biochem 182:319–326

    Article  CAS  PubMed  Google Scholar 

  9. Ton-That H, Liu G, Mazmanian SK et al (1999) Purification and characterization of sortase, the transpeptidase that cleaves surface proteins of Staphylococcus aureus at the LPXTG motif. Proc Natl Acad Sci 96:12424–12429

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgments

The Ministry of Education, Culture, and Science (Gravitation program 024.001.035) is acknowledged for financial support.

Author information

Authors and Affiliations

Authors

Corresponding author

Correspondence to Jan C. M. van Hest .

Editor information

Editors and Affiliations

Rights and permissions

Reprints and permissions

Copyright information

© 2018 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Schoonen, L., van Hest, J.C.M. (2018). Modification of CCMV Nanocages for Enzyme Encapsulation. In: Udit, A. (eds) Protein Scaffolds. Methods in Molecular Biology, vol 1798. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-7893-9_6

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-7893-9_6

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-7892-2

  • Online ISBN: 978-1-4939-7893-9

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics