Recombinant Cercopithecine herpesvirus 1 Envelope glycoprotein D (gD)
Description
Function of Glycoprotein D in Herpesviruses
Glycoprotein D is essential for the entry of alphaherpesviruses into host cells. It binds to specific receptors on the cell surface, which triggers a series of conformational changes in other viral glycoproteins, ultimately leading to the fusion of viral and cellular membranes . In HSV-1, gD binds to HVEM and nectin-1, facilitating viral entry . For B virus, which is closely related to CeHV-1, nectin-1 is a primary receptor, but it does not utilize HVEM efficiently .
Research Findings and Applications
While specific research on recombinant CeHV-1 gD is limited, studies on related viruses like B virus and HSV-1 provide insights into the potential applications of such proteins. These include:
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Vaccine Development: Understanding the role of gD in viral entry can aid in developing vaccines targeting this protein to prevent infection.
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Therapeutic Antibodies: Monoclonal antibodies against gD can be used to neutralize viruses, offering a potential therapeutic approach .
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Diagnostic Tools: Recombinant gD proteins can be used in diagnostic assays to detect antibodies against CeHV-1.
Data Tables
Since specific data on recombinant CeHV-1 gD is not available, we can look at the characteristics of related glycoproteins:
| Glycoprotein | Source | Expression System | Fragment | Tag | Purity |
|---|---|---|---|---|---|
| gI (CeHV-1) | Yeast | His-tag or Tag free | 1-109 aa | His-tag | >90% |
| gE (CeHV-1) | E. coli | His-tag or Tag free | 25-539 aa | His-tag | >90% |
References Glycoprotein D (gD) plays an essential role in cell entry of many simplexviruses. B virus (Macacine herpesvirus 1) is closely related to herpes simplex virus 1 (HSV-1) and encodes gD, which shares more than 70% amino acid similarity with HSV-1 gD . Virus Adsorption And... The results of PCR amplification are shown for the virus isolate BV-ΔgDZ, which was selected and plaque purified on VD60 cells and repassaged three times on Vero cells . Recombinant Cercopithecine herpesvirus 1 (CeHV-1) Envelope glycoprotein I (1-109 aa) was expressed in Yeast with a His-tag or Tag free . Recombinant Cercopithecine herpesvirus 1 (CeHV-1) Envelope glycoprotein E (25-539 aa) was expressed in E. coli with a His-tag or Tag free . B virus and HSV are homologous α-herpesviruses with 57% gD identity, and studies have demonstrated that Nectin-1 can also mediate fusion between the gD of B virus and cells .
Product Specs
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Target Background
Q&A
What is Cercopithecine herpesvirus 1 and why is glycoprotein D significant?
Cercopithecine herpesvirus 1, commonly known as B virus, is an alphaherpesvirus that naturally infects macaque monkeys. It poses significant health risks to humans, potentially causing fatal encephalomyelitis when transmitted from macaques . Glycoprotein D (gD) is one of the major envelope glycoproteins of this virus and serves as a highly antigenic protein that elicits strong antibody responses. Its significance lies in its utility as a safer alternative for detecting B virus infection without requiring propagation of live virus in biosafety level 4 facilities .
How is recombinant gD of Cercopithecine herpesvirus 1 typically expressed?
Recombinant gD of Cercopithecine herpesvirus 1 can be expressed through several experimental systems:
The most documented approach involves cloning the gD gene into a mammalian expression vector (such as pcDNA3.1) and transfecting it into COS7 cells. Expression can be confirmed using indirect immunofluorescence assay or radioimmunoprecipitation analysis (RIPA) .
What are the optimal detection methods for analyzing recombinant gD-antibody interactions?
Research indicates varying effectiveness among detection methods for recombinant gD-antibody interactions:
How does modification of the gD structure affect its utility in diagnostic applications?
Researchers have explored structural modifications of gD to improve its diagnostic utility. A significant advancement was the development of a mutant gD protein lacking the transmembrane domain (TM) and cytoplasmic tail (CT). This modification creates a secretory form of the protein with several advantages:
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The modified protein is secreted into culture medium, simplifying purification
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It maintains proper antigenicity without apparent loss of epitope recognition
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It shows reduced nonspecific reactions in dot blot analysis
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It eliminates the need for cell lysis and subsequent purification steps
This secretory form has proven particularly effective for dot blot analysis, where sera from B virus-infected monkeys react specifically with the protein without the nonspecific reactions sometimes observed with the full-length version .
How can researchers address contradictory results when analyzing gD-antibody interactions?
When faced with contradictory results in gD research, such as variable reactivity across different detection methods, researchers should consider several factors:
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Detection method influences: Different methods (RIPA vs. WB) preserve antigenic epitopes differently, potentially explaining disparate results .
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Data analysis perspective: Following principles from scientific data interpretation, researchers should examine data from multiple angles. As noted in one study, "in the evolution of real knowledge, [contradiction] marks the first step in progress" .
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Sample conditions: Variations in sample preparation, antibody titers, or timing of sample collection relative to infection can affect results.
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Controls: Implementing appropriate positive and negative controls helps distinguish true biological variability from technical artifacts.
A systematic approach to these contradictions often leads to deeper insights. Rather than dismissing contradictory results, researchers should embrace them as potential indicators of complex biological phenomena that warrant further investigation .
What essential controls should be included in serological assays using recombinant gD?
To ensure reliable and interpretable results when using recombinant gD in serological assays, researchers should include:
In published research, sera from monkeys naturally infected with B virus (confirmed positive by ELISA using inactivated B virus antigen) successfully precipitated gD in RIPA, while no specific bands were detected with sera from uninfected monkeys . This clear differentiation between positive and negative samples is essential for assay validation.
How do antibody responses to recombinant gD correlate with natural B virus infection?
Antibody responses to recombinant gD show strong correlation with natural B virus infection, making it a valuable diagnostic tool. In experimental studies, the expressed gD was specifically precipitated with sera from monkeys that had been confirmed to contain antibodies against B virus by ELISA using inactivated virus antigen .
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All positive sera tested showed reactivity in RIPA
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Only some positive sera showed strong reactivity in Western blotting
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The secretory form of gD showed good reactivity in dot blot analysis
These observations suggest that while antibody responses to recombinant gD correlate with infection, the detection method significantly influences the apparent strength of this correlation.
What expression system should researchers choose for optimal recombinant gD production?
The choice of expression system for recombinant gD production depends on research goals and available resources:
For applications requiring the most native-like protein conformation, mammalian expression in COS7 cells has been well-validated. The gene encoding gD can be cloned into pcDNA3.1(-) and transfected into COS7 cells, resulting in expression of immunologically reactive protein .
How can researchers interpret non-specific reactions in immunoassays using recombinant gD?
Non-specific reactions in immunoassays using recombinant gD present a significant challenge for researchers. One study reported nonspecific bands with mobility similar to that of gD when some monkey sera were reacted with extracts from cells transfected with both the gD-expressing plasmid and vector DNA .
To address this issue:
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Compare reactions with vector-only controls to identify non-specific binding
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Use the secretory form of gD (lacking TM and CT) which has been shown to reduce nonspecific reactions in dot blot analysis
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Implement background subtraction based on negative control reactions
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Consider pre-absorption of sera with non-infected cell lysates to reduce background
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Optimize blocking conditions to minimize non-specific interactions
The source of non-specific reactions may include antibodies against cellular proteins, cross-reactivity with related viral antigens, or interactions with the expression system components .
How might contradictions in experimental results with recombinant gD lead to new research insights?
Contradictions in experimental results, rather than being obstacles, can serve as catalysts for new research insights. As noted in one study, "In formal logic, a contradiction is the signal of defeat, but in the evolution of real knowledge, it marks the first step in progress" .
For recombinant gD research, several contradictions merit further investigation:
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Variable reactivity in different immunoassay formats: Understanding why sera that react strongly in RIPA show reduced reactivity in Western blotting could reveal important information about epitope conformation and antibody binding.
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Differential reactivity among positive sera: Exploring why some B virus-positive sera react differently with recombinant gD could uncover variations in immune responses or virus strains.
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Effects of structural modifications: Further characterizing how deletion of the TM and CT domains preserves antigenicity while reducing background could inform the design of improved diagnostic antigens.
By embracing these contradictions as research opportunities, investigators can develop more sophisticated understanding of B virus immunology and improve diagnostic approaches .
What methodological advancements could improve recombinant gD-based diagnostic applications?
Several methodological advancements could enhance the utility of recombinant gD for B virus diagnostics:
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Epitope mapping to identify immunodominant regions that could be expressed as smaller, more stable peptides with retained antigenicity
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Development of multiplex assays incorporating multiple B virus glycoproteins to increase sensitivity and specificity
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Standardization of expression and purification protocols to reduce batch-to-batch variability
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Adaptation of the secretory gD system to high-throughput formats for large-scale surveillance studies
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Integration with modern immunoassay platforms such as bead-based multiplex systems or automated ELISA workstations
These advancements would build upon the established utility of recombinant gD while addressing current limitations in sensitivity, specificity, and scalability for diagnostic applications .
