Fig. 2
Technical route for constructing L1-Env nanoparticles. HPV-16 L1 proteins were expressed in E. coli and subsequently assembled into L1-VLPs in vitro. The L1-VLPs were incubated with TFP-Ester-PEG4-DBCO to attach DBCO to the particle surface, allowing conjugation via copper-free click chemistry. Aldehyde-labeled Env trimers were expressed in HEK293F cells while co-expressing FGE to enhance the conversion of cysteine to formylglycine. Subsequently, the trimers were ligated with aminooxy-PEG3-azide to label them with reactive azide groups at the C-terminus. Finally, the DBCO-labeled VLPs and the azide-labeled Env trimers were conjugated via click chemistry.
