Recombinant Human WSC domain-containing protein 1 (WSCD1)

What is WSCD1 and what are its key structural features?

WSCD1 (WSC domain containing 1) is a protein encoded by the WSCD1 gene located on chromosome 17p13.2 in humans . The protein contains a WSC domain, which is typically associated with carbohydrate binding functions in various proteins. As a recombinant protein, WSCD1 can be produced in various expression systems including mammalian cells (HEK293), E. coli, and other systems to facilitate research applications . The protein structure includes regions that confer its sulfotransferase activity, allowing it to catalyze the transfer of sulfate groups to various substrates . Understanding these structural elements is essential for designing experiments to probe WSCD1 function in cellular pathways and disease mechanisms.

What are the primary biological functions associated with WSCD1?

Based on biochemical characterization, WSCD1 exhibits sulfotransferase activity, suggesting its involvement in post-translational modification of proteins or other biomolecules . This enzymatic function places WSCD1 among other sulfotransferases that participate in various cellular processes including signaling, metabolism, and detoxification. The protein may interact with several other proteins as part of its biological function, though specific pathway associations require further investigation . To study these functions, researchers should design experiments that can directly measure sulfotransferase activity using appropriate substrates and detection methods. When examining WSCD1 in cellular contexts, consideration should be given to tissue specificity and potential regulatory mechanisms that govern its activity.

How is WSCD1 expression regulated in different cell types and tissues?

While the search results don't provide specific information about WSCD1 tissue distribution, researchers interested in expression patterns should employ a systematic approach similar to that used for other proteins like R-Spondin 1, which shows differential expression across tissues . Methodologically, researchers should:

• Utilize quantitative PCR to assess WSCD1 mRNA levels across tissue panels

• Employ immunohistochemistry with validated antibodies to detect protein expression in tissues

• Analyze publicly available RNA-seq datasets to compare expression across cell types and conditions

• Consider single-cell sequencing data to detect cell-type specific expression patterns

These approaches can establish baseline expression patterns that inform experimental design when studying WSCD1 function in specific biological contexts.

What expression systems are optimal for producing functional recombinant WSCD1?

Several expression systems are available for recombinant WSCD1 production, each with distinct advantages for different research applications :

When selecting an expression system, researchers should consider:

• The intended experimental application (structural vs. functional studies)

• Required post-translational modifications for activity

• Protein solubility and stability requirements

• Scale of production needed

For studies investigating WSCD1 sulfotransferase activity, mammalian or insect cell expression systems are generally preferred to maintain native enzymatic function and proper folding .

How should researchers design experiments to study WSCD1 interactions with other proteins?

When investigating WSCD1 protein-protein interactions, researchers should employ a multi-method approach similar to experimental designs used for other recombinant proteins :

• Begin with bioinformatic predictions of potential interaction partners based on domain structure and known interactors

• Perform co-immunoprecipitation experiments with tagged WSCD1 expressed in relevant cell types

• Validate interactions using complementary techniques such as:

  • Pull-down assays with purified recombinant proteins

  • Proximity labeling approaches (BioID or APEX)

  • Yeast two-hybrid screening

  • Surface plasmon resonance for quantitative binding analysis

For data analysis, follow the three-table approach recommended for genomics collaborations :

• First table: Raw interaction data with rows representing potential interactors

• Second table: Sample information including experimental conditions

• Third table: Annotation data providing context for identified interactors

This structured approach ensures comprehensive documentation of experimental parameters and facilitates reproducible analysis of interaction data.

What controls are essential for validating WSCD1 sulfotransferase activity in biochemical assays?

When designing assays to measure WSCD1 sulfotransferase activity, researchers must include appropriate controls to ensure data reliability:

• Negative controls:

  • Heat-inactivated WSCD1 protein

  • Catalytically inactive WSCD1 mutant (site-directed mutagenesis of active site)

  • Reaction mixture without donor substrate (PAPS/APS)

• Positive controls:

  • Well-characterized sulfotransferase with similar substrate preference

  • Commercial sulfotransferase standards with known activity units

• Specificity controls:

  • Substrate competition assays

  • Inhibitor studies with pan-sulfotransferase inhibitors

  • pH and ion concentration gradients to determine optimal conditions

Experimental design should include concentration-dependent measurements to establish Michaelis-Menten kinetics (Km and Vmax values), similar to approaches used for characterizing other enzymes . Activity should be normalized to protein concentration and presented as specific activity (nmol substrate converted/min/mg protein).

How can researchers optimize purification strategies for recombinant WSCD1?

Purification of recombinant WSCD1 should be approached systematically, drawing on principles established for other recombinant proteins :

• Expression optimization:

  • Test multiple expression constructs with different tags (His, Avi, Fc) as indicated in the available forms of WSCD1

  • Optimize induction conditions (temperature, time, inducer concentration)

  • Screen for soluble expression using small-scale pilot experiments

• Purification workflow:

  • Initial capture using affinity chromatography based on the fusion tag

  • Intermediate purification using ion exchange chromatography

  • Polishing step using size exclusion chromatography

• Quality control measures:

  • SDS-PAGE analysis with silver staining to assess purity (aim for >90%)

  • Western blot to confirm identity

  • Mass spectrometry to verify protein integrity and post-translational modifications

  • Dynamic light scattering to assess homogeneity

Monitor protein stability throughout purification using activity assays to ensure the final product retains its sulfotransferase function.

What are the optimal storage conditions for maintaining WSCD1 stability and activity?

While specific stability data for WSCD1 is not provided in the search results, best practices from related recombinant proteins can be applied :

For lyophilized WSCD1, reconstitute in phosphate-buffered saline (PBS) at a concentration of approximately 100 μg/mL, similar to the protocol for R-Spondin 1 . If carrier-free preparations are needed for specific applications, ensure buffers contain stabilizing agents to prevent protein aggregation or activity loss.

Activity testing should be performed after storage periods to confirm retention of sulfotransferase function. Implement a quality control system that tracks protein activity over time under different storage conditions to establish optimal protocols for your specific WSCD1 preparation.

How can researchers verify the structural integrity of purified WSCD1?

To confirm that recombinant WSCD1 maintains its proper structural conformation after purification:

• Biophysical characterization:

  • Circular dichroism (CD) spectroscopy to assess secondary structure content

  • Intrinsic fluorescence to monitor tertiary structure

  • Thermal shift assays to determine stability under different buffer conditions

  • Size exclusion chromatography with multi-angle light scattering (SEC-MALS) to confirm oligomerization state

• Functional verification:

  • Enzyme activity assays to confirm sulfotransferase function

  • Binding assays with known interaction partners

  • Limited proteolysis to assess domain organization and stability

• Structural analysis:

  • For advanced studies, consider X-ray crystallography or cryo-EM

  • Hydrogen-deuterium exchange mass spectrometry for conformational dynamics

Compare multiple production batches to establish consistency metrics and ensure reproducibility in downstream applications. Document all quality control data according to scientific reporting standards .

How can WSCD1 be used in pathway analysis studies?

Given WSCD1's sulfotransferase activity and potential involvement in cellular pathways , researchers can:

• Design pathway perturbation experiments:

  • Overexpress wild-type or mutant WSCD1 in relevant cell models

  • Use CRISPR-Cas9 to knockout or knockdown endogenous WSCD1

  • Apply recombinant WSCD1 exogenously to cellular systems when appropriate

• Perform integrated omics analysis:

  • Phosphoproteomics to identify changes in signaling pathways

  • Metabolomics to detect altered sulfation patterns

  • Transcriptomics to identify gene expression changes

• Data analysis approach:

  • Implement the three-table data organization system for experimental data :

    • Table 1: Primary experimental data (e.g., expression changes)

    • Table 2: Sample information documenting experimental conditions

    • Table 3: Annotation data connecting molecular changes to pathways

• Visualization and interpretation:

  • Use pathway enrichment analysis to identify affected biological processes

  • Build interaction networks incorporating known WSCD1 binding partners

  • Apply statistical methods appropriate for multi-omics data integration

This systematic approach allows researchers to place WSCD1 function within broader cellular contexts and generate testable hypotheses about its role in normal physiology and disease states.

What are the current methodologies for studying WSCD1 in disease models?

Researchers investigating WSCD1's potential roles in disease should consider these methodological approaches:

• Selection of appropriate disease models:

  • Cell lines representing disease-relevant tissues

  • Primary patient-derived cells where appropriate

  • Animal models with WSCD1 mutations or altered expression

  • Organoid systems to better recapitulate tissue architecture

• Functional assessment techniques:

  • Compare WSCD1 expression between normal and disease tissues

  • Analyze substrate sulfation patterns in disease contexts

  • Conduct rescue experiments with recombinant WSCD1 in deficient models

  • Perform structure-function studies with domain mutants

• Experimental design considerations:

  • Include appropriate statistical power calculations

  • Design experiments with proper controls and replication

  • Consider genetic and environmental variables that might influence results

  • Document all experimental conditions thoroughly

• Data collection and analysis:

  • Maintain detailed laboratory records

  • Use standardized reporting formats

  • Apply appropriate statistical tests for the experimental design

  • Consider both parametric and non-parametric approaches based on data distribution

These approaches should be tailored to the specific disease context being investigated, with careful attention to model validity and translational relevance.

How can researchers design experiments to investigate post-translational modifications of WSCD1?

To study post-translational modifications (PTMs) of WSCD1 itself:

• PTM identification strategies:

  • Mass spectrometry-based proteomics with enrichment for specific modifications

  • Site-directed mutagenesis of potential modification sites

  • Antibody-based detection with modification-specific antibodies

  • Chemical labeling approaches for specific PTMs

• Functional impact assessment:

  • Compare activity of modified versus unmodified WSCD1

  • Create phosphomimetic or non-phosphorylatable mutants

  • Assess protein-protein interactions with PTM-specific interactors

  • Determine subcellular localization changes upon modification

• Temporal dynamics investigation:

  • Pulse-chase experiments to track PTM acquisition and turnover

  • Stimulus-response studies to identify regulatory events

  • Cell-cycle synchronization to detect cell-cycle dependent modifications

• Data analysis framework:

  • Document all experimental parameters in the three-table format

  • Apply appropriate normalization to account for expression differences

  • Use statistical methods suited for time-series data when appropriate

  • Visualize results using heat maps or network diagrams

This multi-faceted approach will provide insights into how WSCD1 itself is regulated post-translationally, which may inform its role in cellular pathways and potential as a therapeutic target.

What statistical approaches are recommended for analyzing WSCD1 functional data?

When analyzing data from WSCD1 functional studies, researchers should apply rigorous statistical methods as outlined in experimental design resources :

How should researchers interpret contradictory results in WSCD1 studies?

When faced with contradictory findings about WSCD1 function or interactions:

• Systematic discrepancy analysis:

  • Compare experimental conditions in detail (buffers, tags, expression systems)

  • Evaluate differences in protein preparation methods

  • Consider cell type or tissue-specific effects

  • Examine differences in detection methods or assay sensitivity

• Validation strategies:

  • Replicate findings using multiple independent methods

  • Vary experimental conditions systematically to identify critical parameters

  • Collaborate with other laboratories for independent verification

  • Design critical experiments that can differentiate between competing hypotheses

• Integrated data interpretation:

  • Weigh evidence based on methodological rigor

  • Consider biological context and relevance

  • Evaluate consistency with established mechanisms

  • Acknowledge limitations of each experimental approach

• Transparent reporting:

  • Document all experimental conditions thoroughly

  • Disclose negative and contradictory results

  • Discuss alternative interpretations of the data

  • Suggest definitive experiments to resolve discrepancies

What approaches can researchers use to integrate WSCD1 data into broader -omics datasets?

For integrating WSCD1-specific data with larger -omics datasets:

• Data integration framework:

  • Normalize data across platforms using appropriate methods

  • Apply dimension reduction techniques for visualization (PCA, t-SNE, UMAP)

  • Use correlation analyses to identify relationships between datasets

  • Implement network analysis approaches to place WSCD1 in biological context

• Pathway enrichment approaches:

  • Gene set enrichment analysis (GSEA) using established pathway databases

  • Network-based enrichment to identify functional modules

  • Protein-protein interaction analysis incorporating WSCD1 interactors

  • Text mining of literature to build knowledge networks

• Multi-omics integration strategies:

  • Apply multi-block statistical methods for data fusion

  • Use supervised integration approaches for phenotype association

  • Implement Bayesian networks for causal relationship inference

  • Develop visualization tools that represent multiple data types

• Interpretation guidelines:

  • Distinguish correlation from causation

  • Consider biological plausibility of identified associations

  • Validate key findings with targeted experiments

  • Document computational methods thoroughly for reproducibility

Follow the organizational principles outlined in the three-tables approach, ensuring proper annotation of all experimental metadata . This structured approach facilitates data sharing and collaborative analysis across research groups studying WSCD1 and related proteins.

What are common pitfalls in recombinant WSCD1 expression and purification?

Researchers working with recombinant WSCD1 may encounter several challenges that require specific troubleshooting strategies:

• Low expression yield:

  • Optimize codon usage for the expression host

  • Test different fusion tags and their positions (N- or C-terminal)

  • Adjust induction conditions (temperature, duration, inducer concentration)

  • Consider co-expression with chaperones to improve folding

• Protein insolubility:

  • Test expression at lower temperatures (16-20°C)

  • Screen various buffer compositions during lysis and purification

  • Consider adding solubility enhancers (arginine, glycerol, mild detergents)

  • Explore refolding protocols if inclusion bodies form

• Loss of activity during purification:

  • Include protease inhibitors throughout the purification process

  • Minimize exposure to extreme pH or temperatures

  • Add stabilizing agents like those used for other recombinant proteins

  • Consider carrier proteins like BSA for dilute solutions

• Protein aggregation:

  • Perform buffer optimization screening

  • Use dynamic light scattering to monitor aggregation state

  • Consider addition of non-ionic detergents at low concentrations

  • Implement size exclusion chromatography as a final polishing step

Document all optimization steps systematically to establish a reproducible protocol for future WSCD1 preparations.

How can researchers address non-specific binding in WSCD1 interaction studies?

When investigating WSCD1 protein interactions:

• Sources of non-specific binding:

  • Inappropriate buffer conditions promoting hydrophobic interactions

  • Improper blocking in pull-down or immunoprecipitation experiments

  • Non-native conformations of recombinant proteins

  • Sticky tags or fusion partners

• Experimental controls to implement:

  • Include appropriate negative controls (unrelated proteins with similar properties)

  • Perform competition assays with unlabeled protein

  • Use mutant versions of WSCD1 lacking predicted interaction domains

  • Include detergents or higher salt concentrations to reduce non-specific interactions

• Validation approaches:

  • Confirm interactions using multiple independent methods

  • Perform reciprocal pull-downs (bait-prey reversal)

  • Quantify binding under different conditions

  • Map interaction domains through truncation or mutation analysis

• Data analysis considerations:

  • Establish clear thresholds for distinguishing specific from non-specific interactions

  • Use quantitative measures rather than binary (yes/no) classification

  • Consider relative enrichment compared to control conditions

  • Apply appropriate statistical tests to determine significance

These strategies help distinguish genuine WSCD1 interaction partners from experimental artifacts, increasing confidence in identified protein-protein interactions.

What techniques can overcome challenges in measuring WSCD1 enzymatic activity?

For researchers facing difficulties in detecting or quantifying WSCD1 sulfotransferase activity:

• Sensitivity enhancement strategies:

  • Use radioisotope-labeled substrates (35S-PAPS) for maximum sensitivity

  • Implement coupled enzyme assays to amplify signal

  • Develop fluorogenic substrates for continuous monitoring

  • Consider HPLC or mass spectrometry-based detection for complex substrates

• Substrate identification approaches:

  • Screen candidate substrates based on sequence similarity to known sulfotransferase targets

  • Perform substrate depletion assays with potential substrates

  • Use proteomics to identify sulfated proteins in systems with WSCD1 overexpression

  • Develop in silico prediction methods based on known sulfotransferase preferences

• Assay optimization parameters:

  • Systematic buffer optimization (pH, ionic strength, cofactors)

  • Temperature and time-course studies to determine optimal conditions

  • Enzyme and substrate concentration optimization

  • Addition of stabilizing agents to maintain activity

• Control experiments:

  • Include enzymatically dead mutants as negative controls

  • Use known sulfotransferases as positive controls

  • Incorporate specific inhibitors to confirm signal specificity

  • Perform substrate competition assays to verify binding site interactions

These technical approaches can overcome challenges in detecting and characterizing WSCD1 enzymatic activity, facilitating investigation of its biological function and potential roles in disease mechanisms.

What emerging technologies show promise for advancing WSCD1 research?

Several cutting-edge technologies offer new opportunities for investigating WSCD1:

• CRISPR-based approaches:

  • CRISPR activation/repression for endogenous expression modulation

  • Base editing for introducing specific mutations

  • CRISPR screens to identify genetic interactions with WSCD1

  • Prime editing for precise genomic modifications

• Advanced structural biology techniques:

  • Cryo-electron microscopy for high-resolution structure determination

  • Hydrogen-deuterium exchange mass spectrometry for conformational dynamics

  • Single-molecule FRET for studying conformational changes during catalysis

  • AlphaFold2 and other AI-based structure prediction tools

• Spatial biology methods:

  • Spatial transcriptomics to map WSCD1 expression in tissue context

  • Multiplexed ion beam imaging for protein localization studies

  • Live-cell imaging with fluorescent tags to track dynamics

  • Proximity labeling (BioID, APEX) for spatial interactome mapping

• Single-cell technologies:

  • Single-cell RNA-seq to identify cell populations expressing WSCD1

  • Single-cell proteomics for protein-level analysis

  • Cellular indexing of transcriptomes and epitopes (CITE-seq) for surface expression

Researchers should consider how these emerging technologies might be applied to address specific questions about WSCD1 function, regulation, and role in cellular processes.

How can computational approaches enhance experimental studies of WSCD1?

Computational methods can complement experimental WSCD1 research:

• Structural bioinformatics:

  • Homology modeling of WSCD1 based on related proteins

  • Molecular dynamics simulations to study dynamic properties

  • Docking studies to predict substrate binding and specificity

  • Virtual screening for potential inhibitors or activators

• Systems biology approaches:

  • Network analysis to predict WSCD1 pathway involvement

  • Mathematical modeling of enzymatic activity in cellular contexts

  • Machine learning to identify patterns in multi-omics data

  • Flux balance analysis to understand metabolic impact

• Evolutionary analysis:

  • Comparative genomics across species to identify conserved features

  • Phylogenetic analysis to trace evolutionary history

  • Positive selection analysis to identify functionally important residues

  • Coevolution analysis to predict interaction interfaces

• Integrated data analysis:

  • Multi-omics data integration approaches

  • Text mining of scientific literature

  • Development of WSCD1-specific databases or knowledge bases

  • Pathway visualization and analysis tools

These computational approaches should be integrated with experimental validation following the rigorous design principles outlined in experimental design resources , creating a feedback loop between in silico predictions and laboratory verification.

What are the most promising translational applications for WSCD1 research?

Based on its sulfotransferase activity and potential involvement in cellular pathways, WSCD1 research may have several translational implications:

• Biomarker development:

  • Expression or activity levels as potential diagnostic markers

  • Post-translational modification patterns as disease indicators

  • Development of assays to measure WSCD1 activity in clinical samples

  • Integration into multi-protein diagnostic panels

• Therapeutic targeting strategies:

  • Small molecule modulators of WSCD1 activity

  • Protein-protein interaction disruptors for specific pathways

  • Gene therapy approaches for conditions with altered WSCD1 function

  • Structure-based drug design targeting WSCD1 active site

• Research tool applications:

  • Development of activity-based probes for sulfotransferase activity

  • Creation of reporter systems for studying WSCD1 regulation

  • Generation of conditional knockout models for tissue-specific studies

  • Engineering of WSCD1 variants with enhanced or altered activities

• Methodological innovations:

  • Novel assay technologies for sulfotransferase activity measurement

  • Improved recombinant protein production methods

  • Advanced imaging approaches for tracking sulfation in living cells

  • High-throughput screening platforms for WSCD1 modulators

Researchers pursuing translational applications should maintain rigorous experimental design principles and consider regulatory and ethical implications for clinical development pathways.

How can researchers effectively organize and share WSCD1 experimental data with collaborators?

To facilitate productive collaborations in WSCD1 research:

• Data organization framework:

  • Implement the three-table approach for genomics collaborations :

    • Table 1: Experimental data (e.g., expression values, activity measurements)

    • Table 2: Sample information (experimental conditions, treatments)

    • Table 3: Feature annotations (gene information, pathway associations)

  • Use standardized file formats (CSV, TSV) with clear headers and documentation

  • Maintain consistent naming conventions across all datasets

• Collaboration platforms:

  • Utilize electronic lab notebooks for real-time sharing of protocols and results

  • Establish shared repositories (GitHub, OSF) for code and data

  • Implement version control for tracking changes to protocols and analyses

  • Use collaborative visualization tools for interactive data exploration

• Documentation standards:

  • Create detailed protocols with all experimental parameters

  • Document all statistical analyses and code with comments

  • Develop metadata standards specific to WSCD1 experiments

  • Maintain an accessible glossary of terms and abbreviations

• Quality control measures:

  • Implement regular data validation checks

  • Cross-validate key findings between laboratories

  • Establish minimum reporting standards for WSCD1 experiments

  • Conduct periodic review of shared datasets for consistency