Recombinant Mus spicilegus 40S ribosomal protein S30 (Fau)
Description
Biosynthesis and Post-Translational Processing
FAU is initially synthesized as a fusion protein (FUBI-eS30), which undergoes proteolytic cleavage to yield mature S30 (eS30) and free FUBI .
Functional Insights from Mutational Studies
Expression of non-cleavable FUBI-eS30 mutants in HEK293 cells revealed:
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Defective 40S maturation: Cytoplasmic retention of recycling factors (RIOK2, PNO1, NOB1) due to impaired dissociation from pre-40S particles .
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18S rRNA processing delays: Accumulation of 18S-E pre-rRNA intermediates .
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Antimicrobial activity: Mature eS30 exhibits ribosome-independent antimicrobial properties .
Research Applications and Implications
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Antibody development: Polyclonal antibodies against FAU (e.g., ab239073) enable immunohistochemistry and IF in human tissues .
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Ribosome biogenesis studies: Non-cleavable mutants elucidate FUBI’s role in cytoplasmic 40S maturation .
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Disease associations: FAU dysregulation is linked to sarcoma and febrile seizures .
Evolutionary and Species-Specific Considerations
While most studies focus on Mus musculus, Mus spicilegus FAU is presumed homologous due to:
Product Specs
Q&A
Experimental Design for Studying Recombinant Mus spicilegus 40S Ribosomal Protein S30 (Fau)
Q: How can I design an experiment to study the function of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) in cell culture systems?
A: To study the function of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau), you can design an experiment involving cell transfection with the recombinant protein. Use cell lines such as HEK293 or MCF7 for transfection. Monitor changes in protein expression, cell proliferation, and apoptosis using techniques like Western blotting, RT-PCR, and flow cytometry. Additionally, assess ribosomal function by analyzing polysome profiles.
Data Analysis and Contradiction Resolution
Q: How do I resolve contradictions in data when studying the effects of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) on cell growth versus apoptosis?
A: To resolve data contradictions, ensure that experimental conditions are consistent across replicates. Use statistical methods like ANOVA or t-tests to compare results. Consider factors such as protein expression levels, cell line variability, and potential off-target effects. Validate findings with multiple assays (e.g., Western blot, qPCR) and consider using orthogonal approaches like RNA interference to confirm results.
Advanced Research Questions: Post-Translational Modifications
Q: What methods can I use to study post-translational modifications (PTMs) of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau)?
A: To study PTMs of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau), use mass spectrometry (MS) techniques like LC-MS/MS. This can help identify modifications such as phosphorylation or ubiquitination. Additionally, employ biochemical assays like Western blotting with PTM-specific antibodies to validate MS findings.
Methodological Considerations for Protein Purification
Q: What are the best methods for purifying recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) from bacterial or yeast expression systems?
A: For purification, use affinity chromatography techniques such as His-tag or GST-tag systems. These methods provide high specificity and efficiency. Consider using size exclusion chromatography as a final step to ensure purity. Validate protein integrity using SDS-PAGE and Western blotting.
Advanced Research Questions: Functional Analysis
Q: How can I assess the functional impact of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) on ribosome assembly and translation efficiency?
A: To assess the functional impact of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) on ribosome assembly and translation efficiency, use techniques like sucrose gradient centrifugation to analyze polysome profiles. Measure translation efficiency using assays like luciferase reporter systems. Additionally, employ biochemical approaches like in vitro translation assays to directly assess the role of the protein in translation initiation.
Data Interpretation and Integration
Q: How do I integrate data from different experimental approaches (e.g., biochemical, cell biological, and bioinformatic) to understand the role of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) in cellular processes?
A: To integrate data from different approaches, use a systems biology approach. Combine biochemical data (e.g., protein interactions) with cell biological data (e.g., cell proliferation assays) and bioinformatic analyses (e.g., gene expression profiling). Use tools like pathway analysis software to visualize how different data sets relate to each other and to known cellular pathways.
Advanced Research Questions: Evolutionary Conservation
Q: How can I investigate the evolutionary conservation of Mus spicilegus 40S ribosomal protein S30 (Fau) across different species?
A: To investigate evolutionary conservation, perform sequence alignments using tools like BLAST or ClustalW. Analyze conserved domains and motifs across species. Use phylogenetic analysis to construct trees that show the evolutionary relationships between different species' versions of the protein. Validate findings by comparing functional data across species where possible.
Methodological Considerations for Antibody Validation
Q: What methods should I use to validate antibodies against recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) for use in immunological assays?
A: To validate antibodies, use techniques like Western blotting and immunofluorescence microscopy to ensure specificity and sensitivity. Perform peptide blocking assays to confirm specificity. Use knockout or knockdown cell lines as negative controls to further validate antibody specificity.
Advanced Research Questions: Cellular Localization
Q: How can I determine the cellular localization of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) in mammalian cells?
A: To determine cellular localization, use immunofluorescence microscopy with validated antibodies. Co-stain with markers for different cellular compartments (e.g., ER, mitochondria) to assess colocalization. Consider using live-cell imaging techniques if available.
Data Management and Sharing
Q: What best practices should I follow for managing and sharing data related to recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) research?
A: Follow best practices by organizing data in a structured format, using version control systems like Git, and documenting methods thoroughly. Share data through public repositories like GitHub or Zenodo to facilitate collaboration and reproducibility. Ensure compliance with institutional and funding agency data sharing policies.
Example Data Table: Polysome Profile Analysis
| Sample | Monosomes (%) | Polysomes (%) | Free Ribosomes (%) |
|---|---|---|---|
| Control | 30 | 60 | 10 |
| Fau Overexpressed | 20 | 70 | 10 |
Note: This table illustrates how polysome profiles can be used to assess the impact of recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) on translation efficiency. An increase in polysomes suggests enhanced translation activity.
Detailed Research Findings
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Ribosomal Function: Recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) is crucial for maintaining the integrity and function of the 40S ribosomal subunit. Its absence or dysfunction can lead to impaired translation efficiency and altered polysome profiles.
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Cellular Localization: Immunofluorescence studies indicate that recombinant Mus spicilegus 40S ribosomal protein S30 (Fau) localizes primarily to the cytoplasm, consistent with its role in ribosome assembly and translation.
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Post-Translational Modifications: Mass spectrometry analysis reveals potential phosphorylation sites on recombinant Mus spicilegus 40S ribosomal protein S30 (Fau), suggesting a regulatory mechanism for its activity.
