Recombinant Human Protein disulfide-isomerase (P4HB) is produced in E. coli and provided as a full-length mature protein spanning amino acids 18-508. The product includes an N-terminal 6xHis-tag for straightforward purification and detection. SDS-PAGE analysis confirms a purity level exceeding 90%, meeting high-quality standards for research applications. Endotoxin levels are kept minimal to support various experimental protocols.
Protein disulfide-isomerase (P4HB) appears to play a central role in disulfide bond formation, which seems essential for proper protein folding. The enzyme catalyzes thiol-disulfide exchange reactions, contributing to protein structural stability. P4HB represents a key component in cellular processes related to protein synthesis and maintenance, making it an important target for studies examining protein folding and cellular homeostasis.
Potential Applications
Note: The applications listed below are based on what we know about this protein's biological functions, published research, and experience from experts in the field. However, we haven't fully tested all of these applications ourselves yet. We'd recommend running some preliminary tests first to make sure they work for your specific research goals.
Human P4HB is an enzyme that requires precise folding, proper disulfide bond formation (with multiple conserved disulfide bonds), and specific active site conformation for its functional activity in catalyzing disulfide bond rearrangement. The E. coli expression system can provide an oxidative environment that supports disulfide bond formation, which is favorable for P4HB folding. However, the N-terminal 6xHis-tag may sterically interfere with the protein's functional domains or active site. While the full-length mature protein (18-508aa) contains all functional domains, the probability of correct folding with full enzymatic activity requires experimental validation. The homologous bacterial expression system increases the likelihood of proper folding, but tag interference remains a concern.
1. Protein Folding and Disulfide Bond Formation Studies
This application carries a significant risk without functional validation. P4HB's enzymatic activity requires precise tertiary structure and active site formation. If correctly folded and active (verified through isomerase assays), the protein is suitable for studying disulfide bond formation. If misfolded/inactive (unverified), experiments will yield biologically meaningless results due to impaired catalytic function.
2. Antibody Development and Immunoassay Applications
This application is highly suitable as antibody development relies on antigenic sequence recognition rather than functional enzymatic activity. The full-length protein provides comprehensive epitope coverage for generating antibodies against P4HB. The high purity (>90%) ensures minimal contamination-related issues.
3. Protein-Protein Interaction Studies
This application requires proper folding validation. P4HB interactions with substrate proteins require precise tertiary structure. If correctly folded (verified), the protein may identify physiological interaction partners. If misfolded/unverified, there is a high risk of non-specific binding or failure to replicate genuine interactions.
4. Structural Biology and Biophysical Analysis
These studies are essential for determining folding status. Techniques should include circular dichroism spectroscopy to assess secondary structure, size-exclusion chromatography to evaluate oligomeric state, and enzymatic assays to validate activity. However, the His-tag may interfere with crystallization for high-resolution structural studies.
Final Recommendation & Action Plan
The E. coli expression system is favorable for producing P4HB due to its ability to support disulfide bond formation, but the N-terminal His-tag may cause steric interference. Begin with Application 4 (Structural and Biophysical Analysis) to assess folding quality through CD spectroscopy, SEC, and validate enzymatic activity using standard disulfide isomerase assays. Applications 1 and 3 require rigorous functional validation before proceeding. Application 2 (antibody development) can proceed immediately. Consider tag removal for critical functional studies to minimize potential interference. Always include appropriate activity controls and validate key findings with native P4HB when possible.
