Recombinant Rat Polypeptide N-acetylgalactosaminyltransferase 13 (Galnt13), partial
Description
Functional Role in O-Glycosylation
Galnt13 is critical for:
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Tn Antigen Synthesis: Transfers GalNAc to serine/threonine, forming immunogenic Tn antigens implicated in cancer metastasis .
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Substrate Specificity: Preferentially glycosulates mucin peptides (e.g., Muc5Ac) and proteoglycans like syndecan-3 (SDC3) .
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Neurological Functions: Highly expressed in rat neuronal cells, contributing to brain-specific glycosylation patterns .
Production and Purification
Table 1: Key Studies on Recombinant Rat Galnt13
Clinical and Industrial Applications
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Cancer Research: Trimeric Tn antigen synthesis by Galnt13 enhances metastatic potential in lung and breast cancers .
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Neurological Studies: Role in neurogenesis via glycosylation of podoplanin (PDPN) .
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Toxicology Models: Used to assess glycosylation disruption under heavy metal exposure .
Limitations and Future Directions
Product Specs
Note: We prioritize shipping the format currently in stock. However, if you have specific format requirements, please indicate them when placing your order. We will fulfill your request accordingly.
Note: All our proteins are shipped with standard blue ice packs. If you require dry ice shipping, please contact us in advance. Additional fees will apply.
Generally, the shelf life of liquid form is 6 months at -20°C/-80°C. Lyophilized form has a shelf life of 12 months at -20°C/-80°C.
Target Background
Q&A
Recombinant Rat Polypeptide N-acetylgalactosaminyltransferase 13 (Galnt13) plays critical roles in O-glycosylation, with implications in cancer biology and cellular regulation. Below are FAQs structured for academic researchers, addressing experimental design, methodological challenges, and advanced research considerations.
How should I design experiments to express and purify recombinant rat Galnt13 for functional studies?
Recombinant Galnt13 requires careful handling due to its enzymatic sensitivity and structural complexity:
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Expression systems: Use mammalian systems (e.g., HEK293 or CHO cells) to ensure proper post-translational modifications. The catalytic domain (Ser29-Thr556) is critical for activity .
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Carrier protein considerations: Opt for carrier-free formulations if downstream applications (e.g., structural studies) require minimal interference, but use BSA-supplemented versions for cell culture or ELISA standards .
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Storage: Maintain at -80°C in Tris-NaCl buffers to prevent aggregation, and avoid repeated freeze-thaw cycles .
What methods validate Galnt13 enzymatic activity in vitro?
Use phosphatase-coupled assays with UDP-GalNAc as a donor substrate and synthetic peptides (e.g., RSLLPALRAVISRNQE) . Key steps:
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Substrate libraries: Employ oriented random peptide substrates (e.g., GAGAXXXXXTXXXXXAGA) to profile catalytic specificity .
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Glycopeptide analysis: Confirm glycosylation sites via MALDI-TOF or surface plasmon resonance (SPR) to quantify binding kinetics .
How do I resolve contradictions in Galnt13’s role across cancer studies?
Galnt13 exhibits context-dependent roles:
Methodological approach:
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Compare isoform-specific splice variants (e.g., ΔEx9, Ex10b) using Western blotting with custom monoclonal antibodies .
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Conduct tissue-specific knockout models to isolate microenvironmental effects .
What technical challenges arise when studying Galnt13 splice variants?
Nine splice variants of Galnt13 have been identified, with functional implications:
| Variant | Domain Alteration | Enzymatic Activity |
|---|---|---|
| Wild-type | Full catalytic/lectin | Active |
| Δ39bpEx9 | Truncated lectin | Inactive |
| Ex10b | Modified lectin | Partially active |
Solutions:
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Use insect cell expression systems (e.g., Sf9) for higher yields of unstable variants .
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Validate activity with glycopeptide libraries (e.g., GP(T22)R vs. GP(T10)L) to assess lectin domain function .
How does Galnt13’s substrate specificity compare to its paralog Galnt1?
Galnt13 and Galnt1 share 88% catalytic domain identity but differ in lectin domain function:
| Feature | Galnt13 | Galnt1 |
|---|---|---|
| Substrate preference | Binds N- and C-terminal glycopeptides equally | Favors C-terminal glycopeptides |
| Tissue expression | Brain-specific | Ubiquitous |
| Cancer relevance | Linked to metastasis | Less studied |
Experimental design:
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Use SPR biosensors to compare binding kinetics of synthetic peptides .
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Profile hierarchical glycosylation patterns in mucin-rich cell lines (e.g., MUC3A) .
Methodological Limitations and Future Directions
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Species discrepancy: Most data derive from human Galnt13; rat homolog studies require cross-reactive antibodies .
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Functional redundancy: Overlapping substrate specificities with Galnt1 complicate isoform-specific analyses .
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Splice variant characterization: Only 3/9 variants have been functionally tested .
