Recombinant Human AP-1 complex subunit beta-1 (AP1B1) is produced in an E. coli expression system, covering the 1-584 amino acid region. This partial protein comes engineered with dual tags: an N-terminal 10xHis-tag and a C-terminal Myc-tag, which makes purification and detection more straightforward. The product reaches a purity exceeding 85%, as verified by SDS-PAGE analysis, ensuring reliable results for research applications.
AP1B1 is a subunit of the adaptor protein complex 1 (AP-1), which appears to play a critical role in mediating protein transport between the trans-Golgi network and endosomes. AP1B1 seems essential for maintaining proper protein sorting and trafficking, contributing to cellular homeostasis and molecular signaling pathways. Studying this protein is vital for understanding intracellular transport mechanisms and related cellular processes.
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 dual-tagged AP1B1 protein with N-terminal His-tag and C-terminal Myc-tag provides versatile options for investigating protein interactions within the AP-1 adaptor complex. The His-tag enables immobilization on nickel-based resins for pull-down experiments to identify binding partners. Meanwhile, the Myc-tag allows for detection and validation of interactions through Western blotting or immunoprecipitation. This recombinant protein can be used to study how AP1B1 associates with other AP-1 subunits (γ, μ1, and σ1) or cargo proteins involved in vesicular trafficking. The partial length construct (1-584aa) may reveal specific binding domains responsible for these interactions, though the truncated nature might limit some functional insights.
2. Antibody Development and Validation
This recombinant AP1B1 protein serves as an ideal immunogen and standard for developing specific antibodies against human AP1B1. The high purity (>85%) and dual-tag system make both antibody generation protocols and subsequent validation assays more manageable. Researchers can use this protein in ELISA-based screening of hybridoma clones or phage display libraries to identify high-affinity antibodies. The Myc and His tags provide internal controls for antibody specificity testing. They also enable comparative analysis between tag-specific and AP1B1-specific binding, which can help identify potential cross-reactivity issues.
3. Biochemical Characterization and Structural Studies
The recombinant AP1B1 protein can be used for detailed biochemical analysis including protein folding studies, stability assays, and biophysical characterization techniques such as dynamic light scattering or analytical ultracentrifugation. The dual-tag system makes protein purification optimization and concentration determination easier for structural biology applications. Researchers can investigate the protein's behavior under various buffer conditions, pH ranges, and salt concentrations to establish optimal conditions for downstream applications. The partial construct may provide insights into specific functional domains within the full-length protein, though it's worth noting that some conformational features might be altered compared to the complete protein.
4. In Vitro Reconstitution Assays for AP-1 Complex Assembly
This recombinant AP1B1 subunit can be used in cell-free reconstitution experiments to study AP-1 complex assembly mechanisms and stoichiometry. By combining this protein with other recombinantly expressed AP-1 subunits, researchers can investigate the sequential assembly process and identify critical binding interfaces. The His-tag enables controlled immobilization for surface plasmon resonance or other label-free binding studies to measure association and dissociation kinetics. Such studies likely provide fundamental insights into the molecular basis of AP-1 complex formation and stability, though the artificial expression system may introduce some limitations in mimicking native cellular conditions.
