Recombinant Escherichia coli Transcriptional regulatory protein PhoP is produced using a yeast expression system, which appears to support proper protein folding and post-translational modifications. The protein comes as a full-length construct featuring an N-terminal 6xHis tag and a C-terminal Myc tag - these tags help with purification and detection processes. SDS-PAGE analysis indicates purity levels exceeding 90%, suggesting it may be well-suited for research applications.
PhoP acts as a transcriptional regulatory protein in Escherichia coli and likely plays an important role in how bacteria respond to environmental changes. Working alongside PhoQ, it forms part of a two-component system that controls genes related to magnesium transport and virulence factors. Scientists often examine PhoP to better understand how bacteria adapt and cause disease, which could have broader implications for antibiotic resistance patterns and microbial ecology.
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.
1. Protein-Protein Interaction Studies Using Pull-Down Assays
The N-terminal 6xHis tag makes it possible to attach the protein to nickel-affinity resins for pull-down experiments aimed at finding PhoP binding partners. Having both the 6xHis and C-terminal Myc tags creates options for detecting and confirming protein capture through Western blotting or immunofluorescence methods. This strategy might help reveal PhoP's regulatory network by uncovering new interacting proteins in bacterial lysates or purified protein collections. The high purity level (>90%) should minimize background noise from contaminating proteins during these interaction studies.
2. Antibody Development and Validation
This recombinant PhoP protein could work as an immunogen for creating polyclonal or monoclonal antibodies that target E. coli PhoP specifically. The substantial purity (>90%) makes it a reasonable candidate for immunization protocols and follow-up antibody testing. The C-terminal Myc tag offers a built-in control for checking antibody specificity, helping researchers separate anti-PhoP responses from anti-tag reactions. Researchers might also find this protein useful as a positive control in different immunoassays when validating new antibodies.
3. Biochemical Characterization and Stability Studies
Having the complete recombinant protein (1-223aa) opens doors for thorough biochemical analysis, including thermal stability testing, pH tolerance measurements, and buffer optimization experiments. Scientists can explore protein folding patterns, aggregation tendencies, and storage requirements using methods like dynamic light scattering, circular dichroism spectroscopy, and analytical ultracentrifugation. The dual tag system helps with protein quantification and tracking across various experimental conditions. These studies may be crucial for determining the best handling practices for future functional assays.
4. ELISA-Based Quantitative Assays
The dual tagging approach makes this protein particularly attractive for creating sandwich ELISA assays - the 6xHis tag can handle capture while the Myc tag takes care of detection. This arrangement should allow for precise measurement of PhoP levels in bacterial extracts or culture media. High purity levels suggest reliable standard curve development for quantitative work. These assays could become valuable research tools for monitoring PhoP expression under different growth conditions or stress scenarios in laboratory settings.
