Structure of O‐glycosidically linked oligosaccharides synthesized by the insect cell line Sf9

Thomsen, Darrell R.; Post, Leonard; Elhammer, Åke P.

doi:10.1002/jcb.240430107

Cited by 63 publications

(30 citation statements)

References 37 publications

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“…Although we have not observed any differences between baculovirus and VEE-expressed Norwalk VLPs (Baric et al, submitted), previous experiments have demonstrated subtle variations between insect and mammalian posttranslational modification patterns of recombinant proteins (33,64). For example, baculovirus-and vaccinia virus-expressed papillomavirus VLPs were recently shown to possess notable variations in phosphorylation and glycosylation properties (19).…”

Section: Fig 5 Serum Igg Boost Responses From Nv1contrasting

confidence: 44%

Systemic, Mucosal, and Heterotypic Immune Induction in Mice Inoculated with Venezuelan Equine Encephalitis Replicons Expressing Norwalk Virus-Like Particles

Harrington¹,

Yount

Johnston³

et al. 2002

J Virol

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Norwalk-like viruses (NLVs) are a diverse group of single-stranded, nonenveloped, positive-polarity RNA viruses and are the leading cause of epidemic acute gastroenteritis in the United States. In this study, the major capsid gene of Norwalk virus, the prototype NLV, has been cloned and expressed in mammalian cells using a Venezuelan equine encephalitis (VEE) replicon expression system. Upon infection of baby hamster kidney (BHK) cells with VEE replicon particles (VRPs), the Norwalk virus capsid proteins self-assemble to generate high titers of Norwalk virus-like particles (VLPs) that are morphologically and antigenically analogous to wild-type Norwalk virus. Mice inoculated subcutaneously with VRPs expressing the Norwalk virus capsid protein (VRP-NV1) developed systemic and mucosal immune responses to Norwalk VLPs, as well as heterotypic antibody responses to the major capsid protein from another genogroup I NLV strain (NCFL) isolated from a recent outbreak. A second Norwalk virus capsid clone (NV2) containing three amino acid codon mutations from the NV1 clone was also expressed using VEE replicons (VRP-NV2), but upon infection of BHK cells failed to confer VLP self-assembly. Mice inoculated with VRP-NV2 elicited reduced systemic and mucosal immune responses to Norwalk VLPs, demonstrating the importance and potential utility of endogenous VLP presentation for maximum immune induction. Inoculation with either VRP-NV1 or VRP-NV2 resulted in serum antibody responses far superior to the induction in mice dosed orally with VLPs that were prepared using the VEE-NV1 replicon construct, a regimen similar to current models for NLV vaccination. Expression of NLV VLPs in mammalian cells offers a powerful approach for the design of novel NLV vaccines, either alone or in combination with current vaccination models.Norwalk-like viruses (NLVs) are the most frequent cause of epidemic gastroenteritis in the United States, infecting an estimated 23 million people each year, and have accounted for as much as 96% of selected outbreaks of acute, nonbacterial gastroenteritis (20,40,47). Persons of all ages are affected, and it is unclear whether infection typically confers long-term immunity (38). NLV outbreaks commonly occur within families, nursing homes, day care centers, schools, hospitals, and in the military (1,20,40,42). Infection with NLVs usually results in an episode of acute, self-limited gastroenteritis, with symptoms rarely more serious than nausea, vomiting, diarrhea, and lowgrade fever (40), and most outbreaks are likely not reported (47). However, using refined molecular detection methodologies, recent studies have extended the public health significance of NLV infections and suggest that NLVs are a common cause of severe diarrhea in children (22,53). Although further studies are necessary, NLVs may cause as much as 20% of endemic diarrheal disease in family settings in developed countries (42).The mortality attributable to NLV infection remains unclear, but fatalities among infected elderly may not be uncommon. It ...

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Section: Fig 5 Serum Igg Boost Responses From Nv1contrasting

confidence: 44%

Systemic, Mucosal, and Heterotypic Immune Induction in Mice Inoculated with Venezuelan Equine Encephalitis Replicons Expressing Norwalk Virus-Like Particles

Harrington¹,

Yount

Johnston³

et al. 2002

J Virol

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“…Lectin analyses also show the presence of O-linked oligosaccharidic chains on recombinant subunits and eLH/CG secreted by Sf9 and Mimic TM cells, with a GalNAcGal disaccharide structure (Legardinier et al, submitted) as previously shown for insect or recombinant glycoproteins expressed in Sf9 and other insect cell lines [28,29].…”

Section: Biochemical Properties Of Recombinant Ecg Expressed In Baculsupporting

confidence: 54%

Involvement of equine chorionic gonadotropin (eCG) carbohydrate side chains in its bioactivity; lessons from recombinant hormone expressed in insect cells

et al. 2005

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“…More recently, 0-linked glycosylation was also reported by several groups (Thomsen et al, 1990, Chen et al, 1991, Wathen et al, 1991, Svoboda et al, 1991.…”

mentioning

confidence: 58%

Expression of human interferon‐α2 in Sf9 cells

Sugyiama

Ahorn²,

Maurer‐Fogy³

et al. 1993

European Journal of Biochemistry

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Human interferon a2 (IFN-a2) was expressed in Spodopteru frugiperdu Sf9 insect cells using the baculovirus expression system. The protein purified by immunoaffinity chromatography exhibited biological activity identical to that of leukocyte-derived 'natural' IFN-a2. However, the protein was found to be heterogeneously glycosylated, partially truncated by proteolysis and partially lacking a disulfide bridge. The major product was shown to be 0-glycosylated at the same position as natural human IFN-a2. Enzymatic cleavage, reverse-phase HPLC peptide mapping and plasma-desorption mass spectroscopy analysis revealed the presence of two types of 0-linked carbohydrates. The major 0-linked carbohydrate was found to be the disaccharide galactosyl(/31-3)-N-acetylgalactosamine, the minor component the monosaccharide N-acetylgalactosamine. No evidence for sialylation was found. The non-glycosylated species representing about 40% of the total material were shown to partially lack the C-terminal three amino acids. In addition an unglycosylated, reduction-sensitive dimer was observed. This was formed due to the lack of the N-terminal cysteine normally forming an intramolecular disulfide bridge. Furthermore, a minor species was identified which contains Cysl and Cys98 in a modified form, thereby hindering the formation of a disulfide bridge between these two residues.

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Structure of O‐glycosidically linked oligosaccharides synthesized by the insect cell line Sf9

Cited by 63 publications

References 37 publications

Systemic, Mucosal, and Heterotypic Immune Induction in Mice Inoculated with Venezuelan Equine Encephalitis Replicons Expressing Norwalk Virus-Like Particles

Systemic, Mucosal, and Heterotypic Immune Induction in Mice Inoculated with Venezuelan Equine Encephalitis Replicons Expressing Norwalk Virus-Like Particles

Involvement of equine chorionic gonadotropin (eCG) carbohydrate side chains in its bioactivity; lessons from recombinant hormone expressed in insect cells

Expression of human interferon‐α2 in Sf9 cells

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