• International Journal of Industrial Entomology and Biomaterials
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• v.35 no.2
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• pp.118-122
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• 2017

Porcine pseudorabies virus (PRV) causes the Aujeszky's disease (AD) which is economically important disease in the swine industry worldwide. Killed or live vaccines have been used to control this disease, but their efficacy and side effects remain problems to be solved. To solve these problems, in this study, production of recombinant PRV glycoprotein gB, gC and gD was investigated in insect expression system. Glycoprotein gB, gC and gD are regarded as the major immunogenic antigens in PRV. Abundant production and immunogenicity of glycoprotein gB, gC and gD were confirmed by SDS-PAGE and Western blot analysis, respectively. Optimal infection dose and time were also determined for the production of each recombinant PRV glycoprotein. Confirmation of glycosylation of recombinant gB, gC and gD suggested their usefulness as antigens for the development of diagnosis kit or vaccines for Aujeszky's disease.

• Journal of fish pathology
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• v.30 no.2
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• pp.71-78
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• 2017

The glycoprotein of novirhabdoviruses is known to play a critical role in the determination of host specificity. Viral hemorrhagic septicemia viruses (VHSVs) in different genotypes have different glycoprotein sequences and show different preferences for specific cell lines. In this study, to know whether the glycoprotein is solely responsible for the host cell preference of VHSV, a recombinant VHSV expressing vesicular stomatitis virus (VSV) glycoprotein instead of VHSV IVa glycoprotein (rVHSV-VSV-G) was generated by reverse genetics and inoculated into several fish cell lines, then, cytopathic effect (CPE) and viral growth caused by rVHSV-VSV-G infection were compared with those caused by rVHSV-wild that was previously generated and has the same genomic sequence with wild-type VHSV except a few nucleotides. The plaque numbers of rVHSV-VSV-G were significantly higher in EPC, BF-2 and GF cells than those of rVHSV-wild. However, in HINAE cells (originated from olive flounder), rVHSV-VSV-G titer was significantly lower than rVHSV-wild titer, and both recombinant VHSVs were not grown well in CHSE-214 cells. Although statistical significances were detected in the titers between rVHSV-wild and rVHSV-VSV-G in several cell lines, the cell line-preference order of rVHSV-VSV-G was not different from that of rVHSV-wild. These results suggest that the replacement of VHSV glycoprotein may not completely change host cell preference, and other regions of VHSV might also involve in the determination of host cell preference.

• IMMUNE NETWORK
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• v.12 no.1
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• pp.8-17
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• 2012

Background: Respiratory syncytial virus (RSV) is a major cause of severe lower respiratory tract diseases in infancy and early childhood. Despite its importance as a pathogen, there is no licensed vaccine against RSV yet. The attachment glycoprotein (G) of RSV is a potentially important target for protective antiviral immune responses. Recombinant baculovirus has been recently emerged as a new vaccine vector, since it has intrinsic immunostimulatory properties and good bio-safety profile. Methods: We have constructed a recombinant baculovirus-based RSV vaccine, Bac-RSV/G, displaying G glycoprotein, and evaluated immunogenicity and protective efficacy by intranasal immunization of BALB/c mice with Bac-RSV/G. Results: Bac-RSV/G efficiently provides protective immunity against RSV challenge. Strong serum IgG and mucosal IgA responses were induced by intranasal immunization with Bac-RSV/G. In addition to humoral immunity, G-specific Th17- as well as Th1-type T-cell responses were detected in the lungs of Bac-RSV/G-immune mice upon RSV challenge. Neither lung eosinophilia nor vaccine-induced weight loss was observed upon Bac-RSV/G immunization and subsequent RSV infection. Conclusion: Our data demonstrate that intranasal administration of baculovirus-based Bac-RSV/G vaccine is efficient for the induction of protection against RSV and represents a promising prophylactic vaccination regimen.

• Applied Chemistry for Engineering
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• v.24 no.1
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• pp.62-66
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• 2013

This study was performed to investigate the physicochemical characteristics of glycoprotein extracted from carrot. We manufactured glycoprotein using natural vegetable raw materials carrots, and then analyzed it's physicochemical properties. The results showed that the nutritional composition of carrot glycoprotein (CG) consisted of 2.35% carbohydrate and 94.2% glycoprotein. There were small amounts of hydroxyproline and glycine known as characteristics of collagen peptide in the amino acid of CG, but there were large amount of glutamic acid and aspartic acid involved in the metabolism of glucose and fat. The calories of CG was 342.1 kcal/100 g and the molecular weight appeared less than 594 Da average molecular weight distribution.

• The Plant Pathology Journal
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• v.15 no.6
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• pp.313-318
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• 1999

Cloning of the 5'untranslated region (5' UTR) and Nterminus of the glycoprotein precursor (G2G1) open reading frame of tomato spotted wilt virus has been problematic, possibly because of the toxicity of a signal peptide at the beginning of th G2G1 protein precursor. The toxicity of the signal peptide to bacterial growth and the reason for the expression of the peptide gene in Escherichia coli were investigated by cloning the 5' UTR and the signal peptide sequence separately. Cells transformed with the plasmid containing both the first 30 amino acids of the glycoprotein and the 5' UTR showed a severe growth inhibition whereas transformants harboring either the plasmid with the signal sequence or the 5'UTR alone did not show any ingibition. An E. coli promoter-like sequence was found in the 5'UTR and tis promoter acivity was confirmed with a promoter-less GUS gene cloned downstream of the 5'UTR. In the cloning of the Tomato spotted wilt virus (TSWV) glycoprotein G2G1 open reading frame all the recovered plasmids contained stop codons in the signal sequence region. However, clones containing no stop codon were recovered when the signal sequence and the 5'UTR were cloned separately.

• Journal of Microbiology and Biotechnology
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• v.27 no.6
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• pp.1098-1105
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• 2017

Vesicular stomatitis virus G glycoprotein (VSV-G) has been widely used for pseudotyping retroviral, lentiviral, and artificial viral vectors. The objective of this study was to establish a potential approach for large-scale production of VSV-G. To this end, VSV-G was cloned with an N-terminal His-tag into Pichia pastoris expression vector pPIC3.5K. Three clones ($Mut^s$) containing the VSV-G expression cassette were identified by PCR. All clones proliferated normally in expansion medium, whereas the proliferation was reduced significantly under induction conditions. VSV-G protein was detected in cell lysates by western blot analysis, and the highest expression level was observed at 96 h post induction. VSV-G could also be obtained from the condition medium of yeast protoplasts. Furthermore, VSV-G could be incorporated into Ad293 cells and was able to induce cell fusion, leading to the transfer of cytoplasmic protein. Finally, VSV-G-mediated DNA transfection was assayed by flow cytometry and luciferase measurement. Incubation of VSV-G lysate with the pGL3-control DNA complex increased the luciferase activity in Ad293 and HeLa cells by about 3-fold. Likewise, incubation of VSV-G lysate with the pCMV-DsRed DNA complex improved the transfection efficiency into Ad293 by 10% and into HeLa cells by about 1-fold. In conclusion, these results demonstrate that VSV-G could be produced from P. pastoris with biofunctionalities, demonstrating that large-scale production of the viral glycoprotein is feasible.

• Journal of Applied Biological Chemistry
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• v.61 no.4
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• pp.379-382
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• 2018

Trafficking process of viral glycoprotein to cell surface results in the syncytium formation when baby hamster kidney (BHK) cells was infected by Newcastle disease virus (NDV). Rhus chinensis gall, well-known as a medicinal plant, inhibited not only syncytium formation, but also trafficking of glycoprotein, hemagglutinin-neuramidase (HN) to the cell-surface. Modification of viral glycoprotein is processed within the endoplasmic reticulum and golgi body during trafficking into surface. R. chinensis gall extracts showed the strong inhibitory activities ($IC_{50}$ $12.5{\mu}g/mL$) against ${\alpha}-glucosidase$, when compared with the ${\beta}-glucosidase$. And this inhibitory activities is increased by the samples in a dose-depedent pattern. These data showed that the extracts of R. chinensis gall inhibited the cell-surface expression of NDV-hemagglutinin-neuramidase glycoprotein without significantly affecting HN glycoprotein synthesis in NDV-infected BHK cells.

• BMB Reports
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• v.34 no.4
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• pp.371-378
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• 2001

The Glycoprotein D (gD) gene of the HSV-1 strain F was cloned, sequenced, recombinated into the HcNPV (Hyphantria cunea nuclear polyhedrosis virus) expression vector and expressed in insect cells. The gD gene was located in the 6.43 kb BamHI fragment of the strainF. The open reading frame (ORF) of the gD gene was 1,185 by and codes 394 amino acid residues. Recombinant baculoviruses, GD-HcNPVs, expressing the gD protein were constructed. Spodoptera frugiperda cells, infected with the recombinant virus, synthesized a matured gX-gD fusion protein with an approximate molecular weight of 54 kDa and secreted the gD proteins into the culture media by an immunoprecipitation assay The fusion gD protein was localized on the membrane of the insect cells, seen by using an immunofluorescence assay The deduced amino acid sequence presents additional characteristics compatible with the structure of a viral glycoprotein: signal peptide, putative glycosylation sites and a long C-terminal transmembrane sequence. These results indicate the utility of the HcNPV-insect cell system for producing and characterizing eukaryotic proteins.

• Korean Journal of Food Science and Technology
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• v.37 no.4
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• pp.624-630
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• 2005

Glycoprotein (60 kDa) isolated from Ficus Carica Linnoeus (FCL glycoprotein) was examined by evaluating its hypolipidemic effects on plasma cholesterol levels and hepatic detoxicant enzyme activities in ICR mice. FCL glycoprotein $(100{\mu}g/mL)$ had strong scavenging activities (38%) against lipid peroxyl radicals. When mice were treated with Triton WR-1339 (400 mg/kg), levels of total cholesterol (TC) and low-density lipoprotein (LDL)-cholesterol in plasma significantly increased by 53.9 and 47.5 mg/dL, respectively, compared to the control, whereas, when pretreated with FCL glycoprotein $(100{\mu}g/mL)$, decreased remarkably by 55.4, and 47,0 mg/dL, compared to Triton WR-1339 treatment alone. Interestingly, high-density lipoprotein (HDL)-cholesterol level did not change. Body and liver weights did not change significantly after Triton WR-1339 treatment in presence of FCL glycoprotein. FCL glycoprotein $(100{\mu}g/mL)$ stimulated activities of antioxidative detoxicant enzymes such as superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GPx), whereas GPx activity significantly increased compared to the control. These results suggest FCL glycoprotein has abilities to scavenge lipid peroxyl radicals, lower plasma lipid levels, and stimulate detoxicant enzyme activity in mouse liver.

• Food Science and Preservation
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• v.21 no.5
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• pp.735-739
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• 2014

Ephedra sinica Stapf, known as a medicinal plant, inhibited not only syncytium formation, but also trafficking of viral glycoprotein, hemagglutinin-neuramidase (HN) to the cell-surface. Trafficking of viral glycoprotein to the surface of infected-cells results in syncytium formation in Newcastle disease virus (NDV)-infected baby hamster kidney (BHK) cells. Viral glycoprotein in the infected-cell is processed within the endoplasmic reticulum during routing into surface. The processing of viral glycoprotein like a N-linked oligosaccharide trimming by ${\alpha}$-glucosidase in cell is necessary for virus infection. Methanol extracts showed inhibitory activities ($IC_{50}$ $15{\mu}g/mL$) against ${\alpha}$-glucosidase. This suggested that E. sinica extracts inhibited the cell-surface expression of NDV-HN glycoprotein without significantly affecting HN glycoprotein synthesis in NDV-infected BHK cells.