Abstract
Chikungunya virus (CHIKV) is a rapidly emerging mosquito-borne virus that causes a severe febrile illness with long-lasting arthralgia in humans. As there is no vaccine to protect humans and limit CHIKV epidemics, the virus continues to be a global public health concern. The CHIKV envelope glycoproteins E1 and E2 are important immunogens; therefore, the aim of this study is to produce trimeric CHIKV spikes in insect cells using the baculovirus expression system. The CHIKV E1 and E2 ectodomains were covalently coupled by a flexible linker that replaces the 6K transmembrane protein. The C-terminal E1 transmembrane was replaced by a Strep-tag II for the purification of secreted spikes from the culture fluid. After production in Sf9 suspension cells (product yields of 5.8–7.6 mg/L), the CHIKV spikes were purified by Strep-Tactin affinity chromatography, which successfully cleared the co-produced baculoviruses. Bis(sulfosuccinimidyl)suberate cross-linking demonstrated that the spikes are secreted as trimers. PNGase F treatment showed that the spikes are glycosylated. LC–MS/MS-based glycoproteomic analysis confirmed the glycosylation and revealed that the majority are of the mannose- or hybrid-type N-glycans and <2% have complex-type N-glycans. The LC –MS/MS analysis also revealed three O-glycosylation sites in E1. In conclusion, the trimeric, glycosylated CHIKV spikes have been successfully produced in insect cells and are now available for vaccination studies.
Highlights
Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes a severe febrile illness in humans
Further expansion of the endemic region is expected because of the increasing geographical distribution of mosquito vectors that are competent for CHIKV transmission
We describe the development of a trimeric CHIKV spike vaccine antigen using baculovirus expression vector system (BEVS) in insect cells
Summary
Chikungunya virus (CHIKV) is a mosquito-borne alphavirus that causes a severe febrile illness in humans. The disease is characterized by an acute stage of rash, high fever, and potentially chronic polyarthralgia [1]. As the chronic stage can persist for weeks or even years, the disease has a serious social and economic impact on patients. Over the last few years, the number of CHIKV patients increased as disease outbreaks expanded across the globe from Africa to Asia, the Americas, and Europe [2]. Further expansion of the endemic region is expected because of the increasing geographical distribution of mosquito vectors that are competent for CHIKV transmission. The expanding spread and impact of the viral infection highlight the need for an effective CHIKV vaccine. No licensed vaccine is available yet and few are in clinical testing [3,4]
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