Recombinant Human CDC42BPG Protein: Research Guide

Recombinant Human CDC42BPG Protein is a research-grade serine/threonine kinase target involved in CDC42-mediated cytoskeletal regulation, offering scientists a reliable tool for dissecting Rho GTPase downstream signaling pathways.

Product Name

Recombinant Human CDC42BPG Protein

SKU

rp373392

Protein Class

Serine/Threonine Kinase (MRCK family)

Species Origin

Human (Homo sapiens)

Primary Application

Cell signaling research, cytoskeletal dynamics studies, kinase activity assays

Storage Recommendation

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

Biological Background of CDC42BPG

CDC42BPG (CDC42 Binding Protein Kinase Gamma), also known as MRCK gamma (Myotonic Dystrophy Kinase-Related CDC42-Binding Kinase gamma), is a serine/threonine protein kinase that belongs to the AGC kinase superfamily. It is a downstream effector of the small Rho GTPase CDC42 and plays a central role in the regulation of actin cytoskeleton organization and actomyosin contractility.

CDC42BPG contains an N-terminal kinase domain, a coiled-coil region, a CRIB (CDC42/Rac Interactive Binding) motif, and a C-terminal pleckstrin homology (PH) domain. Through its CRIB motif, CDC42BPG directly interacts with GTP-bound, active CDC42, linking extracellular signals to cytoskeletal remodeling. This signaling axis is critically involved in processes such as cell migration, cell polarity establishment, and tissue morphogenesis.

Dysregulation of CDC42BPG and related MRCK family members has been implicated in cancer cell invasion and metastasis, making this protein a relevant target for oncology research. Recombinant Human CDC42BPG Protein (rp373392) enables researchers to conduct biochemical characterization, substrate identification, inhibitor screening, and pathway mapping in a controlled in vitro setting.

Reconstitution, Handling, and Storage Protocol

Proper handling of Recombinant Human CDC42BPG Protein is essential to preserve its structural integrity and biological activity. Follow the guidelines below to ensure optimal experimental results.

Reconstitution

If the protein is supplied in lyophilized form, briefly centrifuge the vial before opening to collect the lyophilized material at the bottom. Reconstitute by adding the appropriate volume of ultrapure, sterile water or the buffer specified in the product datasheet. Gently pipette to dissolve; do not vortex vigorously, as this may denature the protein. Allow the solution to stand at room temperature for 10–15 minutes to ensure complete dissolution.

Working Aliquots

Once reconstituted, divide the protein into single-use aliquots at volumes appropriate for your experimental needs. This practice minimizes the number of freeze-thaw cycles experienced by any given aliquot, preserving kinase activity over time.

Storage Conditions

Store lyophilized protein at -20°C or -80°C as indicated on the product datasheet. Reconstituted protein should be stored at -80°C. A carrier protein such as 0.1% BSA (bovine serum albumin) may be added to working aliquots to enhance protein stability during long-term storage. Avoid repeated freeze-thaw cycles, as each cycle can lead to aggregation and loss of enzymatic activity.

Thawing

Thaw aliquots gently on ice or at 4°C. Do not thaw at 37°C, as elevated temperatures may accelerate protein degradation. Use immediately after thawing for best results.

Applications in Research

Recombinant Human CDC42BPG Protein (rp373392) is a versatile reagent suitable for a range of biochemical and cell biological applications:

• Kinase Activity Assays: CDC42BPG can be used as an active enzyme in in vitro kinase assays to measure phosphorylation of specific substrates, such as myosin light chain (MLC) and related cytoskeletal effectors. This enables quantitative assessment of kinase activity and evaluation of small-molecule inhibitors.
• Inhibitor Screening: The recombinant protein is suitable for high-throughput or targeted screening of potential CDC42BPG inhibitors, supporting drug discovery programs focused on the MRCK family kinases.
• Protein–Protein Interaction Studies: As a CDC42 effector, this protein can be used in pull-down assays, co-immunoprecipitation studies, and surface plasmon resonance (SPR) experiments to characterize interactions with CDC42 and other binding partners.
• Antibody Validation: Recombinant CDC42BPG serves as a positive control antigen for validating CDC42BPG-specific antibodies in Western blot, ELISA, and immunoprecipitation applications.
• Structural Studies: The purified protein may be used in biophysical studies such as circular dichroism (CD) spectroscopy or differential scanning fluorimetry (DSF/thermal shift assay) to assess protein folding and thermal stability.

Quality Considerations and Troubleshooting

To achieve reproducible and reliable results with Recombinant Human CDC42BPG Protein, researchers should consider the following quality and troubleshooting points:

Purity and Integrity Verification

Before use, verify protein purity by SDS-PAGE and confirm the expected molecular weight band. If the product datasheet specifies purity greater than a stated threshold (e.g., >85% or >95%), ensure that observed purity matches specification. Unexpected bands may indicate degradation or contaminants; contact Aladdin Scientific technical support if discrepancies are observed.

Activity Verification

If kinase activity is lower than expected, consider the following: (1) ensure the protein has not undergone excessive freeze-thaw cycles; (2) verify that the assay buffer conditions (pH, salt concentration, Mg²⁺/ATP concentrations) are appropriate for serine/threonine kinase activity; (3) confirm that the substrate used is a validated CDC42BPG substrate.

Aggregation

Protein aggregation can reduce apparent activity and may lead to variable results. If aggregation is suspected, centrifuge the reconstituted protein at low speed (e.g., 10,000 × g for 5 minutes at 4°C) prior to use to pellet any aggregated material. Use only the clear supernatant.

Lot-to-Lot Consistency

For comparative studies spanning multiple experiments, record the lot number of each vial used. Contact Aladdin Scientific to obtain Certificate of Analysis (CoA) documentation for each lot to confirm specification compliance.

Frequently asked questions

What is CDC42BPG and why is it relevant to cell biology research?

CDC42BPG (also known as MRCK gamma) is a serine/threonine kinase and downstream effector of the small GTPase CDC42. It regulates actin cytoskeletal organization and actomyosin contractility, making it highly relevant to studies of cell migration, polarity, and cancer cell invasion.

How should I store Recombinant Human CDC42BPG Protein to maintain its activity?

Store lyophilized protein at -20°C or -80°C as directed by the product datasheet. Once reconstituted, aliquot into single-use volumes and store at -80°C. Adding a carrier protein such as 0.1% BSA can improve stability. Avoid repeated freeze-thaw cycles, which can cause aggregation and loss of kinase activity.

What applications is Recombinant Human CDC42BPG Protein (rp373392) suitable for?

This protein is suitable for in vitro kinase activity assays, inhibitor screening, protein–protein interaction studies (e.g., pull-down or SPR assays), antibody validation as a positive control antigen, and structural/biophysical characterization studies.

How do I reconstitute lyophilized Recombinant Human CDC42BPG Protein?

Briefly centrifuge the vial to collect lyophilized material, then add the recommended volume of sterile ultrapure water or specified buffer as indicated on the datasheet. Gently pipette to mix without vortexing, and allow 10–15 minutes at room temperature for complete dissolution before aliquoting and storing.

Can CDC42BPG recombinant protein be used for inhibitor screening assays?

Yes. Recombinant Human CDC42BPG Protein is suitable for targeted or high-throughput inhibitor screening applications. It can be used in biochemical kinase assays to evaluate the potency and selectivity of candidate small-molecule inhibitors targeting the MRCK gamma kinase domain.

References

1. Tan I, Seow KT, Lim L, Leung T. Intermolecular and intramolecular interactions regulate catalytic activity of myotonic dystrophy kinase-related Cdc42-binding kinase alpha. Mol Cell Biol. 2001;21(8):2767–2778.
2. Unbekandt M, Olson MF. The actin-myosin regulatory MRCK kinases: regulation, biological functions and associations with human cancer. J Mol Med. 2014;92(3):217–225.
3. Leung T, Chen XQ, Tan I, Manser E, Lim L. Myotonic dystrophy kinase-related Cdc42-binding kinase acts as a Cdc42 effector in promoting cytoskeletal reorganization. Mol Cell Biol. 1998;18(1):130–140.

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