Recombinant Mouse Lysosomal acid Glucosylceramidase (Gba1) comes from an E.coli expression system and contains the complete mature protein sequence spanning amino acids 20 to 515. The protein carries an N-terminal 6xHis-SUMO tag that appears to improve both solubility and purification efficiency. SDS-PAGE analysis shows the product achieves greater than 85% purity, which should provide consistent results for most research applications.
Lysosomal acid Glucosylceramidase, or Gba1, serves a vital function in lysosomal degradation by breaking down glucosylceramide into glucose and ceramide. This enzymatic process is crucial for proper cellular lipid balance. Given its central role in these pathways, Gba1 represents an important research target for understanding lysosomal disorders and related conditions.
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 mouse Gba1 is a lysosomal enzyme that requires correct folding, disulfide bond formation, glycosylation, and proteolytic processing for its enzymatic activity in glucosylceramide metabolism. E. coli, as a prokaryotic system, lacks the eukaryotic chaperones, glycosylation machinery, and lysosomal proteases necessary for proper Gba1 maturation. The SUMO tag may improve solubility, but it cannot compensate for these fundamental limitations. Without experimental validation (e.g., enzyme activity assays), the protein is highly unlikely to be correctly folded or bioactive. The >85% purity indicates low impurities but does not guarantee functional conformation.
1. Protein-Protein Interaction Studies Using His-Tag Affinity Purification
The His-SUMO tag enables pull-down assays to identify potential binding partners. However, if Gba1 is misfolded (as expected in E. coli), interactions may be non-physiological. The tags themselves may cause artifactual binding. Validate any identified partners using native Gba1 from mouse lysosomes or eukaryotic expression systems.
2. Antibody Development and Validation
This application is suitable. The recombinant Gba1 can serve as an immunogen for generating antibodies against linear epitopes, even if misfolded. The high purity supports consistent immunization. However, antibodies generated may not recognize conformational epitopes of native, properly folded and glycosylated Gba1. Validate antibody specificity against endogenous Gba1 from mouse tissues.
3. Biochemical Characterization and Stability Studies
Suitable for basic biophysical analysis of the recombinant Gba1 protein (e.g., thermal stability, aggregation state). However, data will reflect the properties of the misfolded E. coli product, not native Gba1. Do not extrapolate results to physiological enzyme behavior.
4. Structural Biology Sample Preparation
The 85% purity is insufficient for high-resolution structural studies. Even with further purification, the misfolded protein will not yield meaningful structural insights into native Gba1. The SUMO tag and lack of glycosylation make it unsuitable for crystallography or cryo-EM of the physiological enzyme.
Final Recommendation & Action Plan
This E. coli-expressed Gba1 is unsuitable for functional studies without rigorous validation. First, test enzymatic activity using a fluorogenic substrate (e.g., 4-MUG) to confirm glucocerebrosidase function. If inactive (as expected), limit use to non-functional applications like antibody production, but validate antibodies against native Gba1. For structural or interaction studies, use Gba1 expressed in mammalian or insect cells that support proper folding and glycosylation. Always include appropriate controls with native Gba1 when possible.
