Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:2.5.1.18 (glutathione S-transferase)
22,582 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Our aim was to isolate and characterize white spot syndrome virus (WSSV)-binding proteins from shrimp. After a blot of shrimp hemocyte membrane proteins was overlaid with a recombinant WSSV envelope protein (rVP28), the reactive bands on the blot were detected using anti-VP28 antibody. Among three membrane-associated molecules identified by liquid chromatography-tandem mass spectrometry, there was a 25-kDa protein that bound to both rVP28 and WSSV. Since it had a primary structure with high homology to the small GTP-binding protein Rab7, we named it Penaeus monodon Rab7 (PmRab7). The full-length PmRab7 cDNA was obtained, and results from a glutathione S-transferase pull-down assay confirmed specific binding to rVP28. Reverse transcriptase PCR analysis revealed PmRab7 expression in many tissues, and real-time PCR analysis revealed that expression was constitutive. Binding of PmRab7 to rVP28 or WSSV occurred in a dose-dependent manner and was inhibited by anti-Rab7 antibody. In an in vivo neutralization assay, the number of dead shrimp after challenge with WSSV plus PmRab7 (15%) or WSSV plus anti-Rab7 antibody (5%) was significantly lower than after challenge with WSSV alone (95%). In contrast to the WSSV-injected group, shrimp injected with WSSV plus PmRab7 or WSSV plus anti-Rab7 showed no WSSV-type histopathology. We conclude that PmRab7 is involved in WSSV infection in shrimp. This is the first study to identify a shrimp protein that binds directly to a major viral envelope protein of WSSV.
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PMID:PmRab7 is a VP28-binding protein involved in white spot syndrome virus infection in shrimp. 1704 Dec 24

The truncated fragment M' gene, encoding the exterior of the viral envelope protein of PEDV, was subcloned into prokaryotic expression vector pGEX-6p-1. The recombinant plasmid pGEX-6p-M' was constructed and transformed into E. coli BL21(DE3)pLysS for expression. SDS-PAGE analysis showed recombinant truncated M' protein was highly expressed by pGEX-6p-M' and the product fusion protein GST-M' reached 45% in the total bacteria proteins with the analysis of software AlphaImager2200. The preliminary purified recombinant protein was evaluated for its antigenicity and reactivity through Western blotting and indirect enzyme-linked immunosorbent assay (ELISA) with monoclonal antibody against M protein of PEDV and porcine polyclonal anti-PEDV antiserum as the primary antibody. The results indicated the recombinant truncated M' protein should be candidate as a feasible recombinant diagnostic reagent.
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PMID:High-level prokaryotic expression of envelope exterior of membrane protein of porcine epidemic diarrhea virus. 1747 20

White spot syndrome virus (WSSV) is a large, rod-shaped, enveloped double-stranded DNA virus. In this study, VP38, a viral envelope protein, was expressed as a glutathione S-transferase (GST) fusion protein, and a polyclonal antibody against VP38 was obtained. Far-Western blotting and GST pull-down showed that VP38 interacted directly with VP24, a major WSSV envelope protein. In addition, to delineate the interaction region of VP38 with VP24, GST-VP38n (aa 1-142) and GST-VP38c (aa 143-309) were expressed. The GST pull-down assay revealed that VP38 binds via its C-terminal region to VP24. The result implies that VP38 may participate in the formation of the WSSV envelope.
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PMID:The C-terminal region of envelope protein VP38 from white spot syndrome virus is indispensable for interaction with VP24. 1893 21

Infectious bronchitis virus (IBV), a group 3 coronavirus, produces three proteins (IBV E, IBV 3a, and IBV 3b) from subgenomic mRNA 3 during infection. IBV E, a viral envelope protein, plays a role in virus budding, possibly by altering membrane morphology at the virus assembly site. In addition to this role, IBV E may also function as a viroporin, although no data from infected cells have confirmed this possibility definitively. Conversely, the IBV 3a and IBV 3b proteins are nonstructural proteins. These proteins are dispensable for replication in cell culture, but are thought to be important for infection of the natural host. This chapter details methods for generating and screening antibodies to these gene 3 proteins. Antibodies were raised in rabbits following inoculation with IBV-specific peptides and GST fusion proteins, and were screened by immunofluorescence, radioimmunoprecipitation, and immunoblotting.
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PMID:Generating antibodies to the gene 3 proteins of infectious bronchitis virus. 1905 77

A DENV-2 plasmid named pEII*EIII/NS1*,containing sequences encoding portions of the envelope protein that are potentially involved in the induction of neutralizing antibodies and a portion of the NS1 sequence that is involved in protection, is reported in this work. The synthesized subunit protein was recognized by human sera from infected patients and had the predicted size. The immunogenicity of this construct was evaluated using a mouse model in a prime-boost vaccination approach. The priming was performed using the plasmid pEII*EIII/NS1*, followed by a boost with recombinant full-length GST-E and GST-NS1 fusion proteins. The mice showed specific antibody responses to the E and NS1 proteins, as detected by ELISA, compared to the response of animals vaccinated with the parental plasmid. Interestingly, some animals had neutralizing antibodies. These results show that EII*, EIII and NS1* sequences could be considered for the design ofa recombinant subunit vaccine against dengue disease.
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PMID:A plasmid encoding parts of the dengue virus E and NS1 proteins induces an immune response in a mouse model. 2039 Mar 12

The prevalence of feline foamy virus (FFV, spumaretrovirinae) in naturally infected domestic cats ranges between 30 and 80% FFV positive animals depending on age, sex and geographical region analyzed. Two serotypes have been reported for FFV designated FUV7-like and F17/951-like. Serotype-specific neutralization has been shown to correlate with sequence divergence in the surface (SU) domain of the envelope protein (Env). We analyzed a serum collection of 262 domestic cat sera from Germany using a GST-capture ELISA setup screening for Gag and Bet specific antibodies and identified 39% FFV positive animals. Due to the heterogeneity of the serological samples, cut-offs for Gag and Bet reactivity had to be experimentally determined since application of calculated cut-off values yielded some false-positive results; the new cut-off values turned out to be also fully applicable to a previous study. Using the already established FUV7 ElpSU antigen and the newly cloned and produced F17/951 ElpSU antigen, both consisting of the corresponding ectodomains of the envelope leader protein (Elp) and SU protein, we aimed at the detection of Env-specific antibodies and discrimination between the two known FFV serotypes within the diagnostic FFV ELISA. We validated the ElpSU antigens using cat reference sera of known serotype and screened with this assay domestic cat sera from Germany. Use of the FUV7- and F17/951 ElpSU antigens in ELISA resulted in the detection of Env-specific antibodies in both cat reference sera and sera from domestic cats in Germany, but failed to allow serotyping at the same time.
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PMID:Pattern of seroreactivity against feline foamy virus proteins in domestic cats from Germany. 2172 69

Unfolded protein response (UPR) is a cellular adaptive response which functions to reduce stress caused by misfolded proteins in the endoplasmic reticulum (ER). We and others have previously shown that infection with hepatitis C virus (HCV) or expression of the viral proteins can trigger the UPR. HCV is a single-stranded positive-sense RNA virus causing chronic diseases in humans. Its genome encodes two envelope proteins E1 and E2 that mature in the ER to form non-covalently bound native complex and disulphide-bonded aggregates. Apart from the ER targeting proteins, cytosolic forms have been documented. We have previously shown that the ER-targeting E1 and E2 are capable of eliciting the UPR whereas others have shown that the cytosolic-targeting E2 can bind to the ER stress kinase PERK to dampen the UPR. In this report, we further show that the other envelope protein E1, in its cytosolic form, can also bind PERK and dampen the UPR. Using GST-pulldown assay, we show that E1 binds to the cytoplasmic domain of PERK, suggesting interaction of E1 and PERK takes place in the cytoplasm. Using reporter gene assay and Western blotting, we show that cytosolic E1 can repress UPR-induced BiP and CHOP promoter activity and reduce UPR-induced CHOP expression level. Altogether these results suggest opposing functions of ER- and cytosolic forms of HCV envelope proteins depending on their subcellular localization.
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PMID:Hepatitis C Virus Envelope Protein E1 Binds PERK and Represses the Unfolded Protein Response. 2366 8

Dengue virus (DENV; genus Flavivirus) contains a positive-stranded RNA genome. Binding of DENV to host cells is mediated through domain III of the viral envelope protein. Many therapeutic mAbs against domain III have been generated and characterized because of its high antigenicity. We have previously established a novel PCR method named the linear array epitope (LAE) technique for producing monoclone-like polyclonal antibodies. To prove this method could be utilized to produce antibody against epitopes with low antigenicity, a region of 10 aa (V365NIEAEPPFG374) from domain III of the envelope protein in DENV serotype 2 (DENV2) was selected to design the primers for the LAE technique. A DNA fragment encoding 10 directed repeats of these 10 aa for producing the tandem-repeated peptides was obtained and fused with glutathione S-transferase (GST)-containing vector. This fusion protein (GST-Den EIII10-His6) was purified from Escherichia coli and used as antigen for immunizing rabbits to obtain the polyclonal antibody. Furthermore, the EIII antibody could recognize envelope proteins either ectopically overexpressed or synthesized by DENV2 infection using Western blot and immunofluorescence assays. Most importantly, this antibody was also able to detect DENV2 virions by ELISA, and could block viral entry into BHK-21 cells as shown by immunofluorescence and quantitative real-time PCR assays. Taken together, the LAE technique could be applied successfully for the production of antibodies against antigens with low antigenicity, and shows high potential to produce antibodies with good quality for academic research, diagnosis and even therapeutic applications in the future.
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PMID:Production of a neutralizing antibody against envelope protein of dengue virus type 2 using the linear array epitope technique. 2494 92

White spot syndrome virus (WSSV) mainly infects crustaceans through the digestive tract. Whether C-type lectins (CLs), which are important receptors for many viruses, participate in WSSV infection in the shrimp stomach remains unknown. In this study, we orally infected kuruma shrimp Marsupenaeus japonicus to model the natural transmission of WSSV and identified a CL (designated as M. japonicus stomach virus-associated CL [MjsvCL]) that was significantly induced by virus infection in the stomach. Knockdown of MjsvCL expression by RNA interference suppressed the virus replication, whereas exogenous MjsvCL enhanced it. Further analysis by GST pull-down and coimmunoprecipitation showed that MjsvCL could bind to viral protein 28, the most abundant and functionally relevant envelope protein of WSSV. Furthermore, cell-surface calreticulin was identified as a receptor of MjsvCL, and the interaction between these proteins was a determinant for the viral infection-promoting activity of MjsvCL. The MjsvCL-calreticulin pathway facilitated virus entry likely in a cholesterol-dependent manner. This study provides insights into a mechanism by which soluble CLs capture and present virions to the cell-surface receptor to facilitate viral infection.
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PMID:Collaboration between a soluble C-type lectin and calreticulin facilitates white spot syndrome virus infection in shrimp. 2507 Aug 55

White spot syndrome virus (WSSV) is one of the major pathogens of cultured shrimp. Identification of envelope protein interactions has become a central issue for the understanding of WSSV assembly. In this paper, WSSV envelope protein VP52B was fused with GST-tag and expressed in Escherichia coli BL-21(DE3). Immunogold-electron microscopy revealed that VP52B was located on the outside surface of WSSV virions. Far-Western blotting analysis suggested that VP52B might directly interact with a major viral envelope protein VP26, and their interaction was confirmed by GST pull-down assay. Further investigation showed that the VP52B binding domain was located between residues 135-170 of VP26. These findings will enhance our understanding of the molecular mechanisms of WSSV morphogenesis.
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PMID:Characterization of white spot syndrome virus VP52B and its interaction with VP26. 2533 40


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