Synthesis and processing of the rubella virus p110 polyprotein precursor in baculovirus-infected Spodoptera frugiperda cells

Oker‐Blom, Christian; Blomster, Martina; Österblad, Monica; Schmidt, Michel; Åkerman, Karl E.O.; Lindqvist, Christer

doi:10.1016/0168-1702(94)00079-r

Cited by 10 publications

(4 citation statements)

References 30 publications

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“…The glycosylation patterns and resulting size changes found for CHIKV E1 and E2 appear to correspond with the postulated number of glycosylation sites in E1 and E2 (1 and 2, respectively). Our findings are in agreement with results obtained in glycoprotein expression studies for other alphaviruses and for the related Rubellavirus , that used recombinant baculoviruses [26,27,45,46]. …”

Section: Discussionsupporting

confidence: 92%

Functional processing and secretion of Chikungunya virus E1 and E2 glycoproteins in insect cells

Metz

Geertsema

Martina

et al. 2011

Virol J

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BackgroundChikungunya virus (CHIKV) is a mosquito-borne, arthrogenic Alphavirus that causes large epidemics in Africa, South-East Asia and India. Recently, CHIKV has been transmitted to humans in Southern Europe by invading and now established Asian tiger mosquitoes. To study the processing of envelope proteins E1 and E2 and to develop a CHIKV subunit vaccine, C-terminally his-tagged E1 and E2 envelope glycoproteins were produced at high levels in insect cells with baculovirus vectors using their native signal peptides located in CHIKV 6K and E3, respectively.ResultsExpression in the presence of either tunicamycin or furin inhibitor showed that a substantial portion of recombinant intracellular E1 and precursor E3E2 was glycosylated, but that a smaller fraction of E3E2 was processed by furin into mature E3 and E2. Deletion of the C-terminal transmembrane domains of E1 and E2 enabled secretion of furin-cleaved, fully processed E1 and E2 subunits, which could then be efficiently purified from cell culture fluid via metal affinity chromatography. Confocal laser scanning microscopy on living baculovirus-infected Sf21 cells revealed that full-length E1 and E2 translocated to the plasma membrane, suggesting similar posttranslational processing of E1 and E2, as in a natural CHIKV infection. Baculovirus-directed expression of E1 displayed fusogenic activity as concluded from syncytia formation. CHIKV-E2 was able to induce neutralizing antibodies in rabbits.ConclusionsChikungunya virus glycoproteins could be functionally expressed at high levels in insect cells and are properly glycosylated and cleaved by furin. The ability of purified, secreted CHIKV-E2 to induce neutralizing antibodies in rabbits underscores the potential use of E2 in a subunit vaccine to prevent CHIKV infections.

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Section: Discussionsupporting

confidence: 92%

Functional processing and secretion of Chikungunya virus E1 and E2 glycoproteins in insect cells

Metz

Geertsema

Martina

et al. 2011

Virol J

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“…However, both immature (presumably E3E2) and mature, furin-cleaved E2 fractions were found intracellularly, most likely a result of the very high expression levels of the CHIKV proteins. Alphaviral processing intermediates are commonly found in many different expression systems, including recombinant baculoviruses [33], [34], [45], [46]. The triple-banded E2 pattern previously observed upon individual expression of E3E2 [22] was not found after expression using Ac-S27, indicating that in this case all glycoproteins were efficiently glycosylated.…”

Section: Discussionmentioning

confidence: 99%

Effective Chikungunya Virus-like Particle Vaccine Produced in Insect Cells

et al. 2013

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The emerging arthritogenic, mosquito-borne chikungunya virus (CHIKV) causes severe disease in humans and represents a serious public health threat in countries where Aedes spp mosquitoes are present. This study describes for the first time the successful production of CHIKV virus-like particles (VLPs) in insect cells using recombinant baculoviruses. This well-established expression system is rapidly scalable to volumes required for epidemic responses and proved well suited for processing of CHIKV glycoproteins and production of enveloped VLPs. Herein we show that a single immunization with 1 µg of non-adjuvanted CHIKV VLPs induced high titer neutralizing antibody responses and provided complete protection against viraemia and joint inflammation upon challenge with the Réunion Island CHIKV strain in an adult wild-type mouse model of CHIKV disease. CHIKV VLPs produced in insect cells using recombinant baculoviruses thus represents as a new, safe, non-replicating and effective vaccine candidate against CHIKV infections.

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“…It has previously been shown that the baculovirus expression vector system is capable of producing significant amounts of RV-specific proteins (21,23,29) and that recombinant proteins have potential in screening for RV-specific antibodies (12,28). In addition, procaryotic and other eucaryotic systems, as well as synthetic peptide technology, have been used to produce RV-specific antigens in hopes of replacing the expensive and infectious authentic RV antigen (1,9,19,31,34).…”

Section: Discussionmentioning

confidence: 99%

“…In this study, we used the strong polyhedrin gene promoter of Autographa californica nuclear polyhedrosis virus to control expression of the 24S cDNA of RV in S. frugiperda insect cells (33). During infection with this recombinant baculovirus, lepidopteran cell cultures were found to synthesize structural RVspecific protein products that very much resembled those of the authentic RV particle (21). The availability of monoclonal antibodies directed against the E1 protein (12) made it possible to purify the corresponding protein from lysates of recombinant-baculovirus-infected insect cells.…”

Section: Discussionmentioning

confidence: 99%

Detection of rubella virus-specific immunoglobulin M antibodies with a baculovirus-expressed E1 protein

Schmidt

Lindqvist

Salmi

et al. 1996

Clin Diagn Lab Immunol

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The structural proteins of rubella virus (RV) were expressed in insect cells by using the baculovirus expression vector system. The recombinant E1 envelope glycoprotein was purified by immunoaffinity chromatography and used to detect RV-specific immunoglobulin M antibodies in a time-resolved fluoroimmunoassay. Correlation analysis between the reactivities of antibodies against this recombinant E1 and the reactivities against authentic RV antigen shows that purified E1 can detect RV antibodies of the immunoglobulin M type.

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Synthesis and processing of the rubella virus p110 polyprotein precursor in baculovirus-infected Spodoptera frugiperda cells

Cited by 10 publications

References 30 publications

Functional processing and secretion of Chikungunya virus E1 and E2 glycoproteins in insect cells

Functional processing and secretion of Chikungunya virus E1 and E2 glycoproteins in insect cells

Effective Chikungunya Virus-like Particle Vaccine Produced in Insect Cells

Detection of rubella virus-specific immunoglobulin M antibodies with a baculovirus-expressed E1 protein

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