Recombinant Human ATP-dependent RNA helicase DDX3Y is produced in a baculovirus expression system, spanning the full length of the mature protein from amino acids 2 to 660. The protein carries both N-terminal 10xHis tags and C-terminal Myc tags, which makes purification and detection more straightforward. SDS-PAGE analysis shows the product achieves greater than 85% purity, suggesting it should deliver reliable research results.
DDX3Y appears to be an ATP-dependent RNA helicase that participates in various cellular processes. These may include RNA metabolism and translation regulation. The protein seems particularly important for unwinding RNA structures - a process that's essential for RNA processing and transport. Given its involvement in these fundamental pathways, DDX3Y has become a significant research focus for scientists trying to understand cellular RNA dynamics and the mechanisms behind them.
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.
Based on the provided information, the recombinant Human DDX3Y is expressed in a Baculovirus expression system (insect cells), which is a eukaryotic system capable of producing properly folded proteins with post-translational modifications similar to mammalian systems. The protein is expressed as the full-length mature protein (2-660aa), which increases the likelihood of correct folding and functional domain integrity. However, since activity is unverified, the protein cannot be assumed to be correctly folded or bioactive without functional validation. DDX3Y is an ATP-dependent RNA helicase that requires precise folding for its enzymatic activity, including proper ATP-binding and RNA-binding domains. While the baculovirus system is favorable for complex eukaryotic proteins, the presence of dual tags (N-terminal 10xHis and C-terminal Myc) could potentially interfere with protein function. Therefore, while the probability of correct folding is reasonably high, experimental validation is essential to confirm bioactivity.
1. Protein-Protein Interaction Studies Using Pull-Down Assays
The dual tagging system enables affinity purification and detection for pull-down assays. However, if DDX3Y is misfolded, it may not interact authentically with biological partners, leading to non-physiological interactions. The tags facilitate technical execution, but results should be interpreted cautiously until folding is validated. If correctly folded, this application is feasible; otherwise, identified interactions may be artifactual.
2. Antibody Development and Validation
This application is appropriate. The recombinant DDX3Y can serve as an immunogen for antibody generation, as antibodies may recognize linear epitopes even if the protein is misfolded. The dual tags aid purification and detection. However, if misfolded, antibodies may not recognize conformational epitopes of native DDX3Y. Validation against endogenous DDX3Y is recommended to ensure specificity.
3. Biochemical Characterization and Enzyme Kinetics Studies
This application is valuable but highly dependent on correct folding. Techniques for biochemical characterization (e.g., stability assays) can assess folding quality, but enzyme kinetics studies require validated helicase activity. If the protein is misfolded, kinetic data will be invalid. Initial experiments must confirm ATP-dependent RNA unwinding activity before quantitative kinetics.
4. In Vitro Binding Assays and Substrate Specificity Studies
This application is feasible for studying nucleic acid binding properties, as binding may be less conformation-sensitive than enzymatic activity. However, if DDX3Y is severely misfolded, even binding assays may yield misleading results. The tags facilitate detection, but substrate specificity studies should be paired with activity checks to ensure biological relevance.
Final Recommendation & Action Plan
Given the uncertainty in folding and bioactivity, the recommended approach is to first perform biochemical and functional validation. Use techniques like circular dichroism (for secondary structure), size-exclusion chromatography (for oligomeric state), and ATPase/helicase assays (for bioactivity) to confirm proper folding and function. If validated, the protein is suitable for all described applications; if not, focus on antibody development and partial biochemical characterization. Always include appropriate controls (e.g., known substrates/inhibitors for activity assays) and consider using tagged protein constructs in parallel with tag-free versions if tag interference is suspected. The baculovirus expression system is appropriate for this eukaryotic protein, but activity confirmation remains crucial for functional studies.
