Secreted Trimeric Chikungunya Virus Spikes from Insect Cells: Production, Purification, and Glycosylation Status
نویسندگان
چکیده
Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne that causes severe febrile illness with long-lasting arthralgia in humans. As there no vaccine to protect humans and limit CHIKV epidemics, the continues be global public health concern. The envelope glycoproteins E1 E2 are important immunogens; therefore, aim of this study produce trimeric spikes insect cells using baculovirus expression system. ectodomains were covalently coupled by flexible linker replaces 6K transmembrane protein. C-terminal was replaced Strep-tag II for purification secreted from culture fluid. After production Sf9 suspension (product yields 5.8–7.6 mg/L), purified Strep-Tactin affinity chromatography, which successfully cleared co-produced baculoviruses. Bis(sulfosuccinimidyl)suberate cross-linking demonstrated as trimers. PNGase F treatment showed glycosylated. LC–MS/MS-based glycoproteomic analysis confirmed glycosylation revealed majority mannose- or hybrid-type N-glycans <2% have complex-type N-glycans. LC –MS/MS also three O-glycosylation sites E1. In conclusion, trimeric, glycosylated been produced now available vaccination studies.
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ژورنال
عنوان ژورنال: Processes
سال: 2022
ISSN: ['2227-9717']
DOI: https://doi.org/10.3390/pr10010162