Post‐Assembly Modification of Protein Cages by Ubc9‐Mediated Lysine Acylation

Abstract

Although viruses have been successfully repurposed as vaccines, antibiotics, and anticancer therapeutics, they also raise concerns regarding genome integration and immunogenicity. Virus‐like particles and non‐viral protein cages represent a potentially safer alternative but often lack desired functionality. Here, we investigated the utility of a new enzymatic bioconjugation method, called lysine acylation using conjugating enzymes (LACE), to chemoenzymatically modify protein cages. We equipped two structurally distinct protein capsules with a LACE‐reactive peptide tag and demonstrated their modification with diverse ligands. This modular approach combines the advantages of chemical conjugation and genetic fusion and allows for site‐specific modification with recombinant proteins as well as synthetic peptides with facile control of the extent of labeling. This strategy has the potential to fine‐tune protein containers of different shape and size by providing them with new properties that go beyond their biologically native functions.

Engineered Nonviral Protein Cages Modified for MR Imaging

“…AaLS-13 is an evolved variant of the cage-forming enzyme lumazine synthase from Aquifex aeolicus. AaLS-13 self-assembles from 360 monomer proteins into 38 nm icosahedrally symmetric cages (Figure a). − Owing to its negatively supercharged interior and large keyhole-shaped surface pores, AaLS-13 encapsulates positively charged cargo at rates approaching the diffusion limit. , Additionally, the surface-exposed termini of AaLS-13 offer further functionalization opportunities, which have already been exploited to display antibodies or for enzymatic labeling. − …”

“… 62 , 63 Additionally, the surface-exposed termini of AaLS-13 offer further functionalization opportunities, which have already been exploited to display antibodies 64 or for enzymatic labeling. 65 − 67 …”