Recombinant Escherichia coli Lipopolysaccharide export system protein lptA is produced using an E. coli expression system. The protein includes the complete mature sequence spanning residues 28 to 185. For easier purification and detection, it carries an N-terminal 6xHis-SUMO tag. SDS-PAGE analysis shows the protein reaches over 90% purity, which should provide high-quality material for research work.
LptA appears to be a key component in E. coli's lipopolysaccharide (LPS) transport machinery. The protein likely plays an important role in assembling and moving LPS molecules across the bacterial cell envelope—a process that seems critical for outer membrane integrity and function. This makes the protein particularly useful for researchers studying bacterial membrane biology and LPS transport mechanisms.
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.
E. coli LptA is a periplasmic protein that functions as part of the lipopolysaccharide (LPS) transport system, requiring specific interactions with other Lpt proteins (LptC and LptB) and proper folding for its bridging function. The E. coli expression system is homologous for this protein, increasing the probability of correct folding. However, the large N-terminal SUMO tag (∼15 kDa) may sterically interfere with LptA's interaction interfaces, particularly its binding sites for LptC and LptB. While the protein may be soluble and folded at the monomer level, the probability of correct functional activity in LPS transport is moderate but requires experimental validation due to potential tag interference.
1. Protein-Protein Interaction Studies Using Pull-Down Assays
Protein-protein interactions in the Lpt complex require precise interface accessibility that may be compromised by the large N-terminal tag. The large SUMO tag may sterically block LptA's interaction interfaces with LptC and LptB, leading to false-negative results or altered binding specificity. If correctly folded, it could identify physiological partners, but results require validation with tag-free protein and complementary methods like bacterial two-hybrid systems. Any interaction data would be biologically irrelevant without validation using natively folded protein.
2. Antibody Development and Validation
This recombinant LptA serves as an excellent immunogen for generating antibodies against E. coli LptA. The mature protein sequence ensures relevant epitope coverage. The high purity minimizes antibodies against contaminants. However, antibodies may not efficiently recognize conformational epitopes dependent on the native tag-free structure.
3. Biochemical Characterization and Stability Studies
This is the essential first step to assess protein quality. Techniques like size-exclusion chromatography with multi-angle light scattering (SEC-MALS) can determine oligomeric state, while circular dichroism can analyze secondary structure and thermal stability. These studies provide critical data or evaluating the protein's physical properties.
4. In Vitro Reconstitution Assays for LPS Transport Studies
This application is not recommended without rigorous validation. Functional LPS transport requires precise stoichiometric interactions between Lpt components. The SUMO tag will likely sterically hinder proper complex formation, leading to failed reconstitution and misleading results. Tag-free LptA is essential for meaningful transport assays.
Final Recommendation & Action Plan
This SUMO-tagged recombinant LptA is primarily suitable for antibody development and biochemical characterization but requires careful validation for interaction studies and is unsuitable for functional transport assays. The immediate priority is Application 3 (Biochemical Characterization) to assess folding state via SEC-MALS and CD spectroscopy. Application 2 (Antibody Development) can proceed immediately. Application 1 (Interaction Studies) should be approached with caution and include validation with complementary methods. Avoid Application 4 (Transport Studies) entirely due to the high probability of tag interference. For functional LPS transport studies, tag-free LptA expressed in its native context is essential. Consider SUMO protease cleavage to remove the tag for more reliable interaction and functional studies.
