Bioengineering of air-filled protein nanoparticles by genetic and chemical functionalization.

Research output: Contribution to journalArticlepeer-review

5 Scopus citations

Abstract

Background: Various bacteria and archaea, including halophilic archaeon Halobacterium sp. NRC-1 produce gas vesicle nanoparticles (GVNPs), a unique class of stable, air-filled intracellular proteinaceous nanostructures. GVNPs are an attractive tool for biotechnological applications due to their readily production, purification, and unique physical properties. GVNPs are spindle- or cylinder-shaped, typically with a length of 100 nm to 1.5 μm and a width of 30–250 nm. Multiple monomeric subunits of GvpA and GvpC proteins form the GVNP shell, and several additional proteins are required as minor structural or assembly proteins. The haloarchaeal genetic system has been successfully used to produce and bioengineer GVNPs by fusing several foreign proteins with GvpC and has shown various applications, such as biocatalysis, diagnostics, bioimaging, drug delivery, and vaccine development. Results: We demonstrated that native GvpC can be removed in a low salt buffer during the GVNP purification, leaving the GvpA-based GVNP's shell intact and stable under physiological conditions. Here, we report a genetic engineering and chemical modification approach for functionalizing the major GVNP protein, GvpA. This novel approach is based on combinatorial cysteine mutagenesis within GvpA and genetic expansion of the N-terminal and C-terminal regions. Consequently, we generated GvpA single, double, and triple cysteine variant libraries and investigated the impact of mutations on the structure and physical shape of the GVNPs formed. We used a thiol–maleimide chemistry strategy to introduce the biotechnological relevant activity by maleimide-activated streptavidin–biotin and maleimide-activated SpyTag003-SpyCatcher003 mediated functionalization of GVNPs. Conclusion: The merger of these genetic and chemical functionalization approaches significantly extends these novel protein nanomaterials' bioengineering and functionalization potential to assemble catalytically active proteins, biomaterials, and vaccines onto one nanoparticle in a modular fashion.
Original languageEnglish (US)
JournalJournal of Nanobiotechnology
Volume21
Issue number1
DOIs
StatePublished - Mar 25 2023

Bibliographical note

KAUST Repository Item: Exported on 2023-03-28
Acknowledgements: The research reported in this publication was supported by funding from the King Abdullah University of Science and Technology. We thank the research intern, A. Huang, for protein nanoparticle production and purification assistance.

ASJC Scopus subject areas

  • Biomedical Engineering
  • Applied Microbiology and Biotechnology
  • Bioengineering
  • Pharmaceutical Science
  • Molecular Medicine
  • Medicine (miscellaneous)

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