Recombinant Human Neuronal acetylcholine receptor subunit alpha-9 (CHRNA9), biotinylated, is produced in E. coli and spans amino acids 26-237. The protein includes an N-terminal MBP tag and a C-terminal 6xHis-Avi tag, which appears to offer enhanced purification and detection flexibility. This partial-length protein shows a purity greater than 85% as assessed by SDS-PAGE, which should provide reliable performance in most experimental applications.
The neuronal acetylcholine receptor subunit alpha-9 forms part of nicotinic acetylcholine receptors—integral membrane proteins that participate in synaptic transmission. These receptors seem to play a key role in modulating neurotransmitter release. Understanding them may be crucial for exploring cholinergic signaling pathways. Research on CHRNA9 could be significant for investigating synaptic function and its potential implications in neurological studies.
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.
Based on the provided information, the recombinant Human CHRNA9 is expressed in E. coli, a prokaryotic system that is generally unsuitable for producing functional eukaryotic transmembrane proteins like CHRNA9. CHRNA9 is a subunit of a ligand-gated ion channel that requires precise folding, post-translational modifications (e.g., glycosylation), and often assembly with other subunits for bioactivity. E. coli cannot perform eukaryotic-type modifications, and the expression of a partial fragment (26-237aa) lacking transmembrane and intracellular domains further reduces the likelihood of correct folding and function. The presence of multiple tags (N-terminal MBP, C-terminal 6xHis-Avi) and biotinylation may exacerbate folding issues. Since activity is unverified, the protein is highly likely to be misfolded and inactive. Experimental validation is critical, but the expression system and protein characteristics suggest a low probability of bioactivity.
1. Antibody Development and Validation Studies
This biotinylated CHRNA9 protein fragment may serve as an antigen for generating antibodies, as antibodies can recognize linear epitopes even if the protein is misfolded. The tags and biotinylation aid purification and detection in assays like Western blotting. However, if misfolded (as expected), the resulting antibodies may not bind the native, fully folded CHRNA9 in physiological contexts (e.g., in neuronal tissues), limiting their utility for functional studies. Validation against native CHRNA9 is essential.
2. Protein-Protein Interaction Screening
The dual tagging system and biotinylation enable technical feasibility for pull-down assays, but if CHRNA9 is misfolded, it may not interact authentically with biological partners (e.g., other receptor subunits or ligands), leading to non-physiological interactions. The tags themselves could cause artifactual binding. This application should be considered high-risk without prior folding validation. If used, results must be confirmed with alternative methods using a full-length, correctly folded protein.
3. ELISA-Based Binding Assays
The biotinylated protein can be immobilized on streptavidin-coated plates for ELISA, but if misfolded, conformational epitopes may be altered, resulting in inaccurate binding data for antibodies, ligands, or other molecules. The high purity is adequate, but assays may measure non-specific binding rather than native interactions. This application should be approached cautiously, with validation using native CHRNA9 controls.
4. Surface Plasmon Resonance (SPR) Binding Studies
SPR requires a correctly folded protein for meaningful kinetic data. If CHRNA9 is misfolded, immobilization on streptavidin chips may yield stable surfaces, but binding affinities and kinetics measured for ligands (e.g., acetylcholine or drugs) would be invalid and misleading for drug discovery. Without activity verification, SPR studies are not recommended. If pursued, initial activity assays are essential.
Final Recommendation & Action Plan
Given the high probability of misfolding due to E. coli expression and the partial length of this eukaryotic transmembrane protein, it is crucial to avoid functional applications until folding and bioactivity are validated. Prioritize using this reagent for non-functional purposes, such as antibody development for linear epitopes (with the understanding that antibodies may need further validation), and avoid interaction or binding studies until the protein's native conformation is confirmed via circular dichroism, size-exclusion chromatography, or comparison with mammalian-expressed CHRNA9. If functional studies are essential, consider alternative expression systems (e.g., insect or mammalian cells) that support proper folding and modifications. Always include controls with native protein when possible.
