GGCT2;3 Antibody
Description
GGCT2;3: Biochemical Context
GGCT2;3 belongs to the GGCT superfamily, which includes enzymes like γ-glutamyl cyclotransferase (GGCT) in humans and Arabidopsis. These enzymes hydrolyze γ-glutamyl bonds in GSH conjugates, influencing cellular redox balance and detoxification . In Arabidopsis, GGCT2;1 and GGCT2;2 are known to degrade cytosolic GSH, but GGCT2;3’s specific function remains unclear .
Table 1: GGCT Isoforms in Arabidopsis
| Isoform | Function | Stress Response Role |
|---|---|---|
| GGCT2;1 | GSH degradation, arsenic tolerance | Arsenic detoxification |
| GGCT2;2 | GSH metabolism | Limited data |
| GGCT2;3 | Potential GSH turnover | Hypothetical (unstudied) |
GGCT2;3 Antibody: Research Gaps and Hypothetical Applications
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GSH dynamics: Antibodies against GGCT enzymes enable immunoblotting and immunohistochemistry to localize enzyme activity .
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Stress responses: GGCT2;1 antibodies confirmed its role in arsenic tolerance by detecting enzyme expression in Arabidopsis .
A hypothetical GGCT2;3 antibody would likely be used for:
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Protein localization: Tracking GGCT2;3 in subcellular compartments (e.g., cytoplasm, nucleus).
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Enzyme activity assays: Correlating GGCT2;3 expression with GSH levels in stress conditions.
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Functional studies: Knockout or overexpression experiments to elucidate its role in GSH metabolism.
Related Antibody Research in Plants
While GGCT2;3-specific data are absent, antibodies for similar enzymes provide a framework:
Table 2: Antibodies for Plant GGCT Isoforms
| Target | Antibody Type | Application | Source |
|---|---|---|---|
| GGCT2;1 | Polyclonal | Immunoblotting, IHC (arsenic studies) | |
| AIG2A/B | Monoclonal | Immunity regulation (salicylic acid) | |
| GGP1 | Polyclonal | TDSM biosynthesis (plant defense) |
Key Insights:
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AIG2A/B antibodies: Demonstrated that catalytic site mutations (E83A) abrogated function, suggesting GGCT activity is critical for enzyme roles .
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GGCT2;1 antibodies: Revealed tissue-specific expression (e.g., high in roots under arsenic stress) .
Potential Mechanisms of GGCT2;3 in GSH Metabolism
GGCT2;3 may function analogously to GGCT2;1 and GGCT2;2 by:
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Catalyzing γ-glutamyl bond hydrolysis: Liberating amino acids from GSH conjugates.
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Regulating GSH turnover: Balancing redox potential during oxidative stress.
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Interacting with stress pathways: Modulating arsenic tolerance or salicylic acid signaling (as seen in AIG2A/B) .
Research Priorities and Limitations
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Antibody Development:
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Specificity challenges: Distinguish GGCT2;3 from GGCT2;1/2 due to sequence homology.
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Validation: Use knockout mutants for antibody testing.
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Functional Studies:
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Stress experiments: Assess GGCT2;3 expression under cadmium, arsenic, or pathogen exposure.
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Metabolite profiling: Link GGCT2;3 activity to GSH, 5-oxoproline, and amino acid levels.
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Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
Q&A
Here’s a structured FAQ collection for GGCT2;3 antibody research, synthesized from peer-reviewed studies and technical documentation:
How is GGCT2;3 antibody validated for specificity in experimental workflows?
Validation requires:
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Knockdown/overexpression controls: Compare signal intensity in GGCT-depleted vs. overexpressing cell lines (e.g., PC3 prostate cancer cells) using Western blot (WB) .
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Species cross-reactivity testing: Assess reactivity in human, mouse, and rat tissues via immunohistochemistry (IHC), noting cytoplasmic vs. nuclear localization patterns .
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Negative controls: Include secondary antibody-only and isotype-matched IgG controls in IHC/WB .
Key validation data:
| Assay | Human Reactivity | Mouse Reactivity | Recommended Dilution |
|---|---|---|---|
| WB | Confirmed | Not tested | 1:1,000–1:5,000 |
| IHC | Strong | Limited | 1:50–1:200 |
What experimental applications are best suited for GGCT2;3 antibody?
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Functional studies: Investigate GGCT’s role in glutathione homeostasis using siRNA knockdown in Arabidopsis or cancer models .
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Pathway analysis: Pair with phospho-STAT3 (Ser727) and c-Met antibodies to study GGCT-STAT3-c-Met signaling axis .
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Immunohistopathology: Score GGCT expression in tumors using intensity/proportion metrics (e.g., high = score >3) .
Observed discrepancies:
Methodological mitigation:
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Normalize to housekeeping genes: Use β-actin or GAPDH in WB, but avoid tubulin due to reported cross-reactivity .
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Tissue-specific controls: Compare adjacent normal tissue in IHC to account for baseline epithelial expression .
What protocols optimize GGCT2;3 antibody for dual immunofluorescence (IF) and WB?
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IF optimization:
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WB troubleshooting:
How does GGCT2;3 antibody performance vary in glutathione-depleted conditions?
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GSH modulation: Treat cells with buthionine sulfoximine (BSO) to deplete glutathione, then quantify GGCT via WB. Expect increased 5-oxoproline levels via HPLC .
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Stress assays: Combine with heavy metal exposure (e.g., arsenic) to study GGCT’s role in redox balance .
