Recombinant Human CHD8 Protein: Research Guide & Protocol

Recombinant Human CHD8 Protein is a chromodomain helicase DNA-binding remodeling factor central to transcriptional regulation, Wnt signaling modulation, and neurodevelopmental research, including studies on autism spectrum disorder.

Protein Name

Recombinant Human CHD8 Protein

Gene Name

CHD8 (also known as HELSNF1, Duplin)

Protein Family

CHD (Chromodomain Helicase DNA-binding) family, subfamily III

Key Functional Domains

Two chromodomains, SNF2-like ATPase/helicase domain, two SANT domains, BRK domain

Primary Research Applications

Chromatin remodeling assays, transcription regulation studies, Wnt/β-catenin pathway research, autism spectrum disorder (ASD) modeling

SKU

rp367301

Recommended Storage

Store at -80 °C; avoid repeated freeze-thaw cycles

Biological Background and Significance of CHD8

CHD8 (Chromodomain Helicase DNA Binding Protein 8) is a member of the CHD family of ATP-dependent chromatin remodeling enzymes, classified within subfamily III alongside CHD7 and CHD9. The protein is encoded by the CHD8 gene and functions as a large nuclear protein that repositions nucleosomes along DNA to regulate accessibility for transcription factors and other chromatin-associated machinery.

CHD8 contains several conserved structural modules: two N-terminal chromodomains that recognize methylated histone tails, a central SNF2-like ATPase/helicase domain responsible for energy-dependent nucleosome remodeling, two SANT domains that mediate histone and DNA interactions, and a C-terminal BRK (Brahma and Kismet) domain. Together, these domains coordinate CHD8's roles in both gene activation and repression depending on genomic context.

CHD8 is a well-established repressor of Wnt/β-catenin target genes; it binds β-catenin and recruits histone H1 to suppress Wnt-responsive promoters. Additionally, CHD8 interacts with the p53 pathway, suppressing p53-mediated apoptosis during early development. Haploinsufficiency of CHD8 is among the most penetrant single-gene risk factors for autism spectrum disorder (ASD), making CHD8 a major focus of neurodevelopmental and psychiatric genetics research.

Handling, Reconstitution, and Storage Protocol

Proper handling of Recombinant Human CHD8 Protein is essential to maintain enzymatic activity and structural integrity throughout experimentation. Follow the guidelines below to ensure optimal performance.

Upon receipt: Inspect the vial for integrity. Immediately transfer to -80 °C if the protein will not be used within 24 hours. Do not store at -20 °C for extended periods, as this may compromise protein stability.

Reconstitution: Allow the vial to equilibrate to room temperature (18–25 °C) for approximately 10–15 minutes before opening. Reconstitute lyophilized protein in the recommended buffer (typically 50 mM Tris-HCl pH 7.5, 150 mM NaCl, 10% glycerol, 1 mM DTT) to the desired working concentration. Gently swirl or pipette to mix; do not vortex. Centrifuge briefly at low speed to collect the solution.

Aliquoting: Prepare single-use aliquots immediately after reconstitution to avoid repeated freeze-thaw cycles, which can denature the protein and diminish ATPase and helicase activity.

Storage conditions: Store aliquots at -80 °C. For short-term use (up to 1 week), working aliquots may be kept at 4 °C. Adding a carrier protein such as 0.1% BSA can improve stability at lower concentrations.

Note: Specific reconstitution buffer composition, recommended concentration, and excipients should be confirmed against the product datasheet (SKU: rp367301) prior to use.

Research Applications and Experimental Considerations

Recombinant Human CHD8 Protein supports a broad range of biochemical and cell-biological research applications:

• Chromatin remodeling assays: CHD8 can be employed in in vitro nucleosome repositioning experiments using mononucleosomal or polynucleosomal templates. ATPase activity can be assessed by coupling ATP hydrolysis to NADH oxidation in a spectrophotometric assay, or by radiolabeled ATP hydrolysis.
• Protein–protein interaction studies: CHD8 interacts with β-catenin, CTCF, p53, and various histone-modifying complexes. Pull-down, co-immunoprecipitation, and surface plasmon resonance (SPR) assays can be performed using the recombinant protein as bait or analyte.
• DNA-binding and ChIP validation: Recombinant CHD8 can be used in electrophoretic mobility shift assays (EMSA) to study sequence-specific or nucleosome-directed DNA binding, and to validate antibodies intended for chromatin immunoprecipitation (ChIP).
• Wnt/β-catenin pathway research: As a regulator of Wnt target gene expression, CHD8 protein is used in transcriptional reporter assays and in vitro binding studies to dissect its repressive mechanism.
• ASD and neurodevelopmental disease models: Recombinant CHD8 supports biochemical characterization of disease-associated variants identified in genomic studies of ASD patients.

Important considerations: CHD8 is a large protein (>200 kDa); verify SDS-PAGE resolution range and ensure gel systems are appropriate. Confirm batch-specific activity data on the certificate of analysis before designing quantitative assays.

Quality, Purity, and Regulatory Considerations

Recombinant Human CHD8 Protein (SKU: rp367301) is intended for research use only (RUO) and is not approved for diagnostic or therapeutic applications in humans. Researchers should note the following quality and compliance considerations:

Purity: Recombinant proteins are typically verified by SDS-PAGE and Coomassie staining or silver staining. Target purity thresholds and actual lot-specific purity values are documented on the certificate of analysis (CoA). Confirm purity specifications meet the requirements of your intended assay before use.

Biological activity: Activity is commonly verified through ATPase assay or nucleosome remodeling assay. Specific activity values and assay conditions are provided on the CoA. Do not use protein from lots that fail internal activity specifications.

Endotoxin: For cell-based assays, endotoxin levels must be considered. Request endotoxin data from the CoA and ensure levels are within acceptable limits for your cell system (typically <1 EU/µg).

Documentation: Retain the CoA and safety data sheet (SDS) for all protein lots used in publications. Some journals require lot-level quality documentation for recombinant proteins.

All specific purity percentages, activity units, and endotoxin values should be taken from the product-specific CoA for SKU rp367301 rather than from this guideline document.

Frequently asked questions

What is CHD8 and why is it important in research?

CHD8 is an ATP-dependent chromatin remodeling enzyme that regulates gene expression by repositioning nucleosomes, controlling Wnt/β-catenin signaling, and interacting with the p53 pathway. It is one of the most frequently mutated genes in autism spectrum disorder (ASD), making it a critical target for neurodevelopmental disease research.

How should Recombinant Human CHD8 Protein be stored to maintain activity?

Recombinant Human CHD8 Protein should be stored at -80 °C in single-use aliquots to avoid repeated freeze-thaw cycles. For short-term use, working aliquots may be stored at 4 °C for up to one week. Always consult the product-specific certificate of analysis for confirmed storage buffer composition and stability data.

What in vitro assays are compatible with Recombinant Human CHD8 Protein?

Recombinant Human CHD8 Protein is compatible with ATPase activity assays, in vitro nucleosome remodeling assays, EMSA for DNA-binding studies, protein–protein interaction assays (pull-down, co-IP, SPR), and antibody validation for ChIP applications. Researchers studying Wnt signaling can also use it in transcriptional repression assays.

Is Recombinant Human CHD8 Protein suitable for use in cell-based assays?

Recombinant Human CHD8 Protein can be used in cell-based assays when endotoxin levels are within acceptable limits (typically less than 1 EU/µg). Always verify endotoxin content on the certificate of analysis before use in cell culture systems to avoid confounding inflammatory responses.

What molecular weight should I expect for CHD8 on SDS-PAGE?

CHD8 is a large protein with a theoretical molecular weight exceeding 200 kDa based on its amino acid sequence. The apparent molecular weight on SDS-PAGE may differ slightly from the predicted value due to post-translational modifications or anomalous migration. Confirm the expected band position from the product certificate of analysis for SKU rp367301.

References

1. Woodage T, Bhatt MA, Bhatt YK, Collins FS. Characterization of the CHD family of proteins. Proc Natl Acad Sci USA. 1997;94(21):11472-11477.
2. Thompson BA, Tremblay V, Lin G, Bochar DA. CHD8 is an ATP-dependent chromatin remodeling factor that regulates β-catenin target genes. Mol Cell Biol. 2008;28(12):3894-3904.
3. Bernier R, et al. Disruptive CHD8 mutations define a subtype of autism early in development. Cell. 2014;158(2):263-276.
4. Nishiyama M, et al. CHD8 suppresses p53-mediated apoptosis through histone H1 recruitment during early embryogenesis. Nat Cell Biol. 2009;11(2):172-182.

Products are supplied for research and development use only. Not for use in humans, animals, diagnosis, or therapy.