Recombinant Human HPD Protein: Properties & Research Guide

Recombinant Human HPD Protein is a research-grade recombinant form of the human 4-hydroxyphenylpyruvate dioxygenase enzyme, enabling scientists to investigate tyrosine catabolism, hereditary metabolic disorders, and potential therapeutic targets in a controlled in vitro setting.

Gene Name

HPD (also known as HPPD)

Protein Full Name

4-hydroxyphenylpyruvate dioxygenase

Biological Function

Catalyzes the conversion of 4-hydroxyphenylpyruvate to homogentisate in the tyrosine degradation pathway

Species Origin

Human (Homo sapiens)

Research Relevance

Implicated in tyrosinemia type III and alkaptonuria; target for herbicide and drug design

Product SKU

rp370090

Supplier

Aladdin Scientific

Biological Background and Function of HPD

4-Hydroxyphenylpyruvate dioxygenase (HPD, also referred to as HPPD) is a non-heme iron-dependent dioxygenase that plays a central role in the catabolism of the aromatic amino acid tyrosine. The enzyme catalyzes the oxidative decarboxylation and rearrangement of 4-hydroxyphenylpyruvate to homogentisate, an intermediate that is subsequently processed to fumarate and acetoacetate through downstream enzymatic steps.

HPD is expressed primarily in the liver and kidney and requires molecular oxygen and ferrous iron (Fe²⁺) as cofactors for its activity. Because homogentisate is a key branch point metabolite in aromatic amino acid catabolism, defects in HPD function are associated with inherited metabolic diseases. Loss-of-function mutations in the HPD gene are linked to tyrosinemia type III, characterized by elevated plasma tyrosine levels and neurological symptoms. Upstream accumulation of homogentisate due to defects in homogentisate dioxygenase (the next enzyme) can also be studied through HPD activity modulation.

Beyond its role in human metabolism, HPD orthologs are targets for inhibitor-based herbicides (e.g., mesotrione, sulcotrione), making recombinant human HPD a valuable model for comparative enzyme studies and selectivity profiling of HPPD-targeted compounds.

Research Applications of Recombinant Human HPD Protein

Recombinant Human HPD Protein is suitable for a range of in vitro biochemical and biomedical research applications:

• Enzyme Activity Assays: The recombinant protein can be used to measure HPD enzymatic activity in defined reaction conditions, enabling kinetic parameter determination (K_m, V_max) and cofactor dependency studies.
• Inhibitor Screening: Researchers studying HPD inhibitors for therapeutic applications (e.g., nitisinone, used clinically for hereditary tyrosinemia type I) or agrochemical development can use recombinant HPD to evaluate compound potency, selectivity, and mechanism of inhibition.
• Structural and Biophysical Studies: Purified recombinant HPD supports protein–ligand binding assays, thermal stability studies, and serves as antigen for antibody generation or validation.
• Disease Mechanism Research: The protein aids investigation of the molecular basis of HPD-associated inborn errors of metabolism, including tyrosinemia type III, by enabling reconstitution of pathway activity in cell-free systems.
• Drug Target Validation: As HPD is explored as a target for management of certain metabolic disorders, recombinant protein supports target engagement and pharmacodynamic studies.

Handling, Storage, and Reconstitution Guidelines

To maintain optimal activity and integrity of Recombinant Human HPD Protein, follow these general best-practice guidelines. Always refer to the specific product datasheet (SKU: rp370090) for lot-specific conditions, as formulation details may vary.

• Storage Temperature: Recombinant proteins are generally stored at −80 °C for long-term stability. Avoid repeated freeze–thaw cycles, which can lead to protein aggregation and loss of enzymatic activity. Aliquot the protein into single-use volumes upon receipt.
• Reconstitution: If supplied lyophilized, reconstitute with the volume and buffer specified in the product datasheet (typically sterile deionized water or a carrier buffer such as PBS with BSA). Allow the vial to equilibrate to room temperature before opening, and mix gently — do not vortex.
• Working Conditions: HPD is an iron-dependent enzyme; ensure assay buffers are free of chelating agents (e.g., EDTA) that could sequester Fe²⁺ and inhibit activity. Supplement reaction buffers with appropriate Fe²⁺ concentrations if required by the assay protocol.
• Protein Concentration: Determine working concentration by preliminary titration experiments. Use a validated protein quantification method (e.g., BCA assay or absorbance at 280 nm) before each experiment.
• Contamination Prevention: Work under aseptic conditions and use sterile, low-binding tubes and pipette tips to minimize protein adsorption and microbial contamination.

Quality, Purity, and Verification

Recombinant proteins intended for research applications should meet defined quality standards to ensure experimental reproducibility. For Recombinant Human HPD Protein (rp370090) from Aladdin Scientific, researchers should consult the product-specific Certificate of Analysis (CoA) and datasheet for verified specifications, which may include:

• Purity Assessment: Typically evaluated by SDS-PAGE under reducing conditions. High-quality recombinant proteins are generally ≥85–95% pure as assessed by densitometry, though the exact threshold should be confirmed with the product datasheet.
• Identity Confirmation: Western blot using an anti-HPD antibody, or mass spectrometry-based peptide mapping, confirms the identity of the recombinant protein.
• Endotoxin Testing: For applications sensitive to lipopolysaccharide (LPS) contamination, endotoxin levels should be verified using a Limulus Amebocyte Lysate (LAL) assay. Check the CoA for reported endotoxin levels.
• Biological Activity: Where applicable, functional enzyme activity may be confirmed by a substrate conversion assay using 4-hydroxyphenylpyruvate as substrate and monitoring homogentisate production.

Always request the CoA for each lot and archive it for GLP or internal quality records.

Regulatory and Safety Considerations

Recombinant Human HPD Protein is supplied for research use only (RUO) and is not intended for diagnostic, therapeutic, or clinical use in humans or animals. Users should observe the following:

• Safety Classification: As a recombinant human protein, standard laboratory biosafety practices apply. Handle in accordance with your institution's biosafety guidelines and any applicable local regulations. Consult the Safety Data Sheet (SDS) provided by Aladdin Scientific for hazard classification and disposal instructions.
• Disposal: Dispose of protein waste, packaging, and consumables in accordance with institutional biosafety protocols and local environmental regulations.
• Export Controls: Some biological research materials may be subject to export control regulations. Verify compliance with applicable regulations before international shipment or transfer.
• Material Transfer: Use of this product should comply with any material transfer agreements (MTAs) applicable to your institution.

If you have questions about regulatory compliance, ordering, or product specifications, contact Aladdin Scientific's customer service team for assistance.

Frequently asked questions

What is the biological function of the HPD protein?

HPD (4-hydroxyphenylpyruvate dioxygenase) catalyzes the conversion of 4-hydroxyphenylpyruvate to homogentisate, a key step in the tyrosine degradation pathway. It is an iron-dependent dioxygenase requiring Fe²⁺ and molecular oxygen as cofactors. Defects in HPD are associated with tyrosinemia type III, a rare inherited metabolic disorder.

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

The protein should be stored at −80 °C for long-term stability. Repeated freeze–thaw cycles should be avoided, so it is recommended to aliquot the protein into single-use volumes upon receipt. Always refer to the product-specific datasheet (SKU: rp370090) for confirmed storage conditions and buffer formulation.

Can Recombinant Human HPD Protein be used for inhibitor screening?

Yes. Recombinant Human HPD Protein is well suited for in vitro inhibitor screening assays. Researchers can use it to evaluate the potency, selectivity, and mechanism of action of candidate HPPD inhibitors, including both therapeutic compounds (such as nitisinone analogs) and agrochemical candidates. Ensure assay buffers are free of Fe²⁺ chelators such as EDTA to maintain enzyme activity during screening.

Is Recombinant Human HPD Protein suitable for clinical or diagnostic use?

No. Recombinant Human HPD Protein (rp370090) is supplied for research use only (RUO) and is not approved or validated for clinical diagnostics, therapeutic administration, or any in vivo use in humans or animals.

What quality control tests are performed on this recombinant protein?

Quality control typically includes purity assessment by SDS-PAGE, identity confirmation by Western blot or mass spectrometry, and endotoxin testing by LAL assay. Functional activity verification may also be included. For lot-specific QC data, request the Certificate of Analysis (CoA) for SKU rp370090 from Aladdin Scientific.

References

1. Lindblad B, Lindstedt S, Steen G. On the enzymic defects in hereditary tyrosinemia. Proc Natl Acad Sci USA. 1977;74(10):4641–4645.
2. Moran GR. 4-Hydroxyphenylpyruvate dioxygenase. Arch Biochem Biophys. 2005;433(1):117–128.
3. Soulère L, Queneau Y, Doutheau A. An expeditious synthesis of nitisinone (NTBC) and of its analogues. Bioorg Med Chem Lett. 2007;17(7):1964–1967.
4. Ellis MK, Whitfield AC, Gowans LA, et al. Inhibition of 4-hydroxyphenylpyruvate dioxygenase by 2-(2-nitro-4-trifluoromethylbenzoyl)-cyclohexane-1,3-dione and 2-(2-nitro-4-methylsulfonylbenzoyl)-cyclohexane-1,3-dione. Toxicol Appl Pharmacol. 1995;133(1):12–19.

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