Recombinant Bacillus subtilis Endoglucanase (eglS) gets expressed in E. coli and includes the complete mature protein sequence, spanning amino acids 30 to 499. The protein carries both an N-terminal 10xHis-tag and a C-terminal V5-tag, which makes purification and detection more straightforward. SDS-PAGE analysis shows it achieves greater than 85% purity, which appears to provide dependable performance for research work.
Endoglucanase from Bacillus subtilis breaks down cellulose by cutting through complex carbohydrates to produce simpler sugars. This enzyme seems central to several biological processes, particularly those related to biomass conversion and energy production. Studying it may help push forward industrial biotechnology research while deepening our grasp of how microbes degrade cellulose.
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.
Bacillus subtilis Endoglucanase (eglS) is a bacterial enzyme that requires precise folding, proper active site formation, and specific tertiary structure for its functional activity in cellulose degradation. The E. coli expression system is homologous to this bacterial protein, which increases the probability of correct folding. The dual N-terminal 10xHis-tag and C-terminal V5-tag are relatively small (∼1 kDa and ∼1.4 kDa, respectively) compared to the protein size (470 aa, ∼50 kDa), minimizing steric interference. While the full-length mature protein (30-499aa) contains all functional domains, the probability of correct folding with functional enzymatic activity requires experimental validation of endoglucanase activity.
1. Enzyme Kinetics and Biochemical Characterization Studies
This application carries a significant risk without functional validation. Endoglucanase activity requires precise active site formation and proper folding. If correctly folded and active (verified through enzymatic assays with cellulose substrates), the protein is suitable for kinetic studies (Km, Vmax). If misfolded/inactive (unverified), kinetic measurements will yield biologically meaningless results.
2. Protein-Protein Interaction Studies
This application requires proper folding validation. Endoglucanase interactions with partners require native conformation. If correctly folded (verified), the protein may identify physiological interaction partners; if misfolded/unverified, there is a risk of non-specific binding or tag-mediated artefacts.
3. 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 mature protein provides comprehensive epitope coverage for generating eglS-specific antibodies. The high purity (>85%) ensures minimal contamination-related issues during immunization.
4. Structural Biology and Protein Folding Studies
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 functionality. If correctly folded, results are valuable; if misfolded, they characterize the recombinant construct.
Final Recommendation & Action Plan
The E. coli-expressed eglS with small dual tags has a high probability of correct folding due to the homologous expression system, but experimental validation of enzymatic activity is crucial. Begin with Application 4 (Structural Biology and Protein Folding Studies) to assess folding quality through CD spectroscopy, SEC, and validate endoglucanase activity using standard cellulose degradation assays (e.g., with carboxymethyl cellulose or other substrates). Applications 1 and 2 require rigorous functional validation before proceeding. Application 3 (antibody development) can proceed immediately. For reliable eglS research, confirm specific activity and kinetic parameters before functional applications, and consider using refolding protocols if initial validation indicates poor functionality.
