Recombinant Human Chemerin-like receptor 1 (CMKLR1) is produced through an E.coli-based cell-free expression system, covering the complete protein sequence from amino acids 1 to 373. The protein contains an N-terminal 10xHis-tag that makes purification and detection more straightforward, reaching purity levels above 85% when analyzed by SDS-PAGE. With its 7-transmembrane domain structure, the protein appears to maintain characteristics of its natural form. The formulation relies on a Detergent Platform approach to help preserve stability.
Chemerin-like receptor 1 (CMKLR1) functions as a G protein-coupled receptor that regulates immune responses and adipocyte activity. This receptor plays an important role in multiple signaling pathways, affecting processes like chemotaxis and inflammation. Given its involvement in these fundamental biological processes, CMKLR1 has become an attractive research target for scientists studying immune regulation and metabolic disorders.
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.
The human CMKLR1 is a GPCR with seven transmembrane domains that requires precise integration into a lipid bilayer, correct folding, and proper disulfide bond formation for bioactivity (e.g., chemerin binding and signaling). While cell-free systems can improve solubility for membrane proteins compared to traditional E. coli expression, they still lack the native membrane environment and eukaryotic chaperones necessary for proper folding of complex GPCRs. The detergent used for solubilization may not replicate the natural lipid bilayer, potentially leading to non-native conformations. Without experimental validation (e.g., ligand binding assays or structural analysis), the protein cannot be assumed to be correctly folded or bioactive.
1. Membrane Protein Solubilization and Detergent Screening Studies
This application is feasible for studying detergent-solubilized CMKLR1 behavior. The His tag facilitates purification during detergent screening. However, the "proper folding" mentioned cannot be assumed without validation. Data will reflect the protein's state in detergents, not necessarily its native conformation. Use results for solubilization optimization but not for inferring in vivo structure.
2. Antibody Development and Epitope Mapping
This application is suitable. The recombinant CMKLR1 can serve as an immunogen for antibody production against linear epitopes, even if misfolded. The high purity supports consistent immunization. However, antibodies generated may not recognize conformational epitopes of native, membrane-embedded CMKLR1. Validate antibody specificity against full-length CMKLR1 expressed in mammalian cells.
3. Protein-Protein Interaction Studies Using Pull-Down Assays
Use with extreme caution. The His-tag enables pull-down assays, but if CMKLR1 is misfolded, interactions are likely non-physiological. GPCRs require correct conformation for specific interactions with G proteins and other partners. The detergent-solubilized state may cause artifactual binding. Validate any identified interactions using full-length CMKLR1 in membrane preparations or cellular systems.
4. Biochemical Characterization and Stability Analysis
It is suitable for basic biophysical analysis of the detergent-solubilized CMKLR1 protein (e.g., thermal stability, aggregation state). However, data may not reflect native GPCR properties due to potential misfolding and detergent effects. Use techniques like circular dichroism to assess secondary structure, but interpret results cautiously without native conformation validation.
Final Recommendation & Action Plan
Before using this recombinant CMKLR1 for functional studies, validate its folding and ligand-binding capability. First, analyze secondary structure via circular dichroism and check oligomeric state via size-exclusion chromatography. Most critically, perform ligand binding assays using labeled chemerin to confirm bioactivity. If active, proceed with controlled interaction studies; if inactive, limit use to non-functional applications like antibody production (with validation against native CMKLR1). For reliable results, consider expressing CMKLR1 in mammalian or insect cell systems that provide proper membrane environments. Always include appropriate controls with native GPCRs when possible.
