Recombinant Human GPR101 Protein: Preparation & Usage Protocol

Recombinant Human GPR101 Protein is a research-grade recombinant protein targeting the orphan G protein-coupled receptor GPR101, designed to support receptor binding studies, functional assays, and neuroendocrine signaling investigations in human biology research.

Product Name

Recombinant Human GPR101 Protein

SKU

rp369941

Protein Class

Orphan G Protein-Coupled Receptor (Class A GPCR)

Species Origin

Human (Homo sapiens)

Primary Applications

Receptor binding assays, functional GPCR assays, neuroendocrine research

Supplier Reference

absea #PP-4274

Background: GPR101 and Its Biological Significance

GPR101 (G Protein-Coupled Receptor 101) is an orphan member of the Class A (rhodopsin-like) superfamily of G protein-coupled receptors (GPCRs). It is predominantly expressed in the central nervous system, with notable enrichment in hypothalamic and pituitary tissues, suggesting roles in neuroendocrine regulation.

GPR101 has been linked to the regulation of growth hormone (GH) and prolactin secretion from the pituitary gland. Somatic duplications and mutations in the GPR101 gene have been associated with X-linked acrogigantism (XLAG), a rare pediatric syndrome characterized by early-onset gigantism driven by pituitary adenomas. These findings have elevated GPR101 as a potential therapeutic target in pituitary tumour biology and disorders of GH excess.

Because no endogenous ligand has been definitively confirmed for GPR101, it remains classified as an orphan receptor, making recombinant protein tools critical for deorphanization studies, binding screens, and functional characterization.

Reconstitution and Storage Instructions

Follow these steps to correctly reconstitute and store Recombinant Human GPR101 Protein (rp369941) to maintain biological activity and protein integrity:

1. Initial inspection: Upon receipt, inspect the vial for integrity. The protein is typically supplied in lyophilised (freeze-dried) form. Centrifuge the vial briefly (1,000–2,000 × g, 30 seconds) before opening to collect any material from the cap or sides.
2. Reconstitution buffer: Reconstitute the lyophilised protein in sterile, deionised water or the buffer recommended in the accompanying Certificate of Analysis (CoA). A typical starting volume yields a stock concentration of 0.1–1.0 mg/mL. Add reconstitution buffer gently along the vial wall and allow to dissolve at room temperature for 10 minutes without vortexing to avoid protein denaturation.
3. Carrier protein: If the protein will be used at very low concentrations or stored for extended periods, consider adding BSA (0.1% final) or another carrier protein as stabiliser, unless it would interfere with downstream assays.
4. Storage conditions: Aliquot reconstituted protein into single-use volumes to avoid repeated freeze-thaw cycles. Store lyophilised protein at –20 °C or –80 °C. Store reconstituted aliquots at –80 °C for long-term storage or at 4 °C for short-term use (≤1 week). Avoid repeated freeze-thaw cycles (maximum 3 cycles recommended).
5. Sterility: Work under aseptic conditions. Filter sterilise (0.22 µm) if the protein will be used in cell-based assays.

Note: Always refer to the product-specific Certificate of Analysis (CoA) supplied with each lot for precise reconstitution volumes, buffer composition, and protein concentration.

Experimental Applications and Usage Guidelines

Recombinant Human GPR101 Protein is suitable for the following research applications:

• Receptor binding assays: Use as a target protein in competitive or saturation radioligand binding assays to identify and characterise putative GPR101 ligands. Optimise protein concentration and buffer conditions empirically for each assay format.
• Deorphanization screening: Incorporate into high-throughput or fragment-based ligand screening platforms to identify candidate endogenous or synthetic ligands for GPR101.
• Antibody validation: Use as a positive control antigen in Western blot, ELISA, or dot-blot experiments when validating anti-GPR101 antibodies.
• Functional GPCR assays: Where applicable, use reconstituted protein in lipid nanodisc or detergent-solubilised systems to assess G protein coupling (Gs, Gi, Gq) using cAMP accumulation assays, GTPγS binding, or BRET/FRET-based biosensor platforms.
• Structural and biophysical studies: Suitable for use in surface plasmon resonance (SPR), isothermal titration calorimetry (ITC), or thermal shift assays to characterise ligand–receptor interactions.

Working concentration: Optimal working concentrations must be determined experimentally and will vary by assay type. Consult the CoA and primary literature for guidance. Titration experiments are strongly recommended before committing to large-scale assays.

Quality Control and Troubleshooting

Quality control checks to perform upon receipt:

• Verify lot number and expiry date on the vial label against the CoA.
• Confirm protein identity by SDS-PAGE under reducing and non-reducing conditions; the observed molecular weight should correspond to that stated on the CoA.
• For activity-sensitive applications, perform a functional activity assay (e.g., binding assay) using the positive and negative controls provided or referenced in the CoA.

Common issues and troubleshooting:

Issue	Possible Cause	Recommended Action
Protein aggregation or precipitation	Incorrect buffer, vortexing during reconstitution, repeated freeze-thaw	Reconstitute gently; use recommended buffer; avoid vortexing; minimise freeze-thaw cycles
Low or no activity	Protein denaturation; improper storage; degradation	Check storage conditions; verify reconstitution procedure; confirm working concentration
Unexpected band on SDS-PAGE	Glycosylation variants; degradation products; dimer formation	Compare to CoA; run under both reducing and non-reducing conditions
High background in assay	Non-specific binding; carrier protein interference	Optimise blocking conditions; titrate protein concentration; consider alternative carrier

Safety, Handling, and Disposal

Safety precautions: Recombinant Human GPR101 Protein is intended for in vitro research use only. It is not approved for use in humans, animals, or clinical diagnostic procedures. Handle all recombinant proteins as potentially biohazardous materials in accordance with your institution's biosafety guidelines.

• Wear appropriate personal protective equipment (PPE): laboratory coat, nitrile gloves, and eye protection.
• Work in a laminar flow hood when handling the protein for sterile cell-based assays.
• Avoid inhalation of lyophilised powder upon reconstitution.

Disposal: Dispose of all protein-containing materials, vials, and consumables in accordance with applicable local, national, and institutional regulations governing biological and chemical waste. Inactivate protein solutions prior to disposal if required by your institutional biosafety office (e.g., autoclaving at 121 °C for 30 minutes).

Regulatory notice: This product is for research use only (RUO). It is not intended for therapeutic or diagnostic use. Consult the Safety Data Sheet (SDS) for full hazard information.

Frequently asked questions

What is GPR101 and why is it considered an orphan receptor?

GPR101 is a Class A G protein-coupled receptor expressed predominantly in the brain and pituitary. It is classified as an orphan receptor because no endogenous ligand has been definitively identified and confirmed for it to date. Recombinant GPR101 protein is a key tool for deorphanization studies aimed at identifying its natural binding partner.

How should I reconstitute and store Recombinant Human GPR101 Protein (rp369941)?

Reconstitute the lyophilised protein using sterile water or the buffer specified in the CoA. Add buffer gently along the vial wall and allow to dissolve at room temperature for 10 minutes without vortexing. Aliquot into single-use volumes and store at –80 °C for long-term use. Avoid more than 3 freeze-thaw cycles to preserve protein activity.

What assay applications is Recombinant Human GPR101 Protein suitable for?

Recombinant Human GPR101 Protein is suitable for receptor binding assays, ligand deorphanization screens, antibody validation (Western blot, ELISA), functional G protein coupling assays (e.g., cAMP accumulation), and biophysical studies such as SPR or ITC. Optimal concentrations should be determined empirically for each application.

Is Recombinant Human GPR101 Protein approved for clinical or therapeutic use?

No. Recombinant Human GPR101 Protein (rp369941) is strictly for in vitro research use only (RUO). It has not been evaluated or approved for use in humans or animals as a therapeutic or diagnostic agent. Always consult your institutional biosafety and ethics guidelines before experimental use.

What is the clinical relevance of GPR101 research?

GPR101 has been implicated in X-linked acrogigantism (XLAG), a rare pediatric syndrome caused by pituitary adenomas that overproduce growth hormone and prolactin. Duplications of the GPR101 gene locus on the X chromosome are a recognised genetic cause of this condition. Research tools such as recombinant GPR101 protein support investigation of its role in pituitary tumour biology and potential therapeutic targeting.

References

1. Trivellin G, Daly AF, Faucz FR, et al. Gigantism and acromegaly due to Xq26 microduplications and GPR101 mutation. New England Journal of Medicine. 2014;371(25):2363-2374. doi:10.1056/NEJMoa1408028
2. Abboud D, Daly AF, Dupuis N, et al. GPR101 drives growth hormone hypersecretion and gigantism in mice via constitutive activation of Gs and Gq/11. Nature Communications. 2020;11(1):4752. doi:10.1038/s41467-020-18500-x
3. Bates B, Zhang L, Nawoschik S, et al. Characterization of Gpr101 expression and G-protein coupling selectivity. Brain Research. 2006;1087(1):1-14. doi:10.1016/j.brainres.2006.02.123

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