Recombinant Human MDH2 Protein: Properties & Lab Guide

Recombinant Human MDH2 Protein is a mitochondrial malate dehydrogenase enzyme critical to the tricarboxylic acid (TCA) cycle, now available from Aladdin Scientific for use in biochemical assays, metabolic research, and enzyme kinetics studies.

Product Name

Recombinant Human MDH2 Protein

SKU

rp369504

Protein Full Name

Malate Dehydrogenase 2, Mitochondrial (MDH2)

Biological Function

Catalyzes the reversible oxidation of malate to oxaloacetate using NAD+ as cofactor in the TCA cycle

Subcellular Localization

Mitochondrial matrix

Primary Applications

Enzyme activity assays, metabolic pathway studies, protein interaction studies, drug target validation

Supplier

Aladdin Scientific

Biological Background and Function of MDH2

Malate Dehydrogenase 2 (MDH2) is a nuclear-encoded mitochondrial enzyme that plays a central role in cellular energy metabolism. It catalyzes the reversible interconversion of L-malate and oxaloacetate coupled to the reduction or oxidation of NAD⁺/NADH within the mitochondrial matrix.

As a key enzyme of the tricarboxylic acid (TCA) cycle, MDH2 is essential for maintaining the flow of carbon through the cycle and supporting mitochondrial respiration. Beyond the TCA cycle, MDH2 contributes to the malate-aspartate shuttle, a mechanism that transfers reducing equivalents from the cytoplasm into the mitochondria, thereby linking cytosolic glycolysis to oxidative phosphorylation.

MDH2 has also attracted significant research interest as a potential biomarker and therapeutic target in cancer metabolism, given the elevated or dysregulated metabolic activity observed in multiple tumor types. Recombinant Human MDH2 Protein enables researchers to study enzyme kinetics, screen inhibitors, and investigate protein-protein interactions in a controlled biochemical setting.

Reconstitution and Storage Guidelines

Proper reconstitution and storage of Recombinant Human MDH2 Protein are essential to preserve enzymatic activity and ensure reproducible experimental results. Follow the guidelines below when handling this reagent.

Reconstitution: Briefly centrifuge the vial before opening to consolidate the lyophilized material at the bottom. Reconstitute in an appropriate sterile aqueous buffer (such as PBS, pH 7.4, or the buffer recommended in the product datasheet). Add the reconstitution volume slowly and gently mix by pipetting — avoid vigorous vortexing, which may denature the protein. Allow a few minutes at room temperature for complete dissolution.

Short-term storage: Once reconstituted, aliquot the protein immediately to avoid repeated freeze-thaw cycles, which degrade enzymatic activity. Store aliquots at −20 °C or −80 °C depending on the intended time frame of use.

Long-term storage: For long-term storage, the lyophilized form should remain at −20 °C. Adding a carrier protein such as 0.1% BSA to the reconstitution buffer may improve stability for dilute working concentrations.

Note: Specific reconstitution volume, buffer composition, and storage conditions are detailed in the product datasheet provided with SKU rp369504. Always refer to the datasheet as the authoritative source.

Experimental Applications and Assay Protocols

Recombinant Human MDH2 Protein is suitable for a range of biochemical and cell-free experimental applications:

1. Enzyme Activity Assay: MDH2 activity can be measured spectrophotometrically by monitoring the oxidation of NADH at 340 nm in a standard malate dehydrogenase assay. In the reverse (reductive) direction, the decrease in absorbance at 340 nm as NADH is consumed during oxaloacetate reduction is used to calculate specific activity. Ensure reagents (oxaloacetate, NADH, assay buffer) are freshly prepared for optimal results.

2. Inhibitor Screening: The recombinant protein is suitable for small-molecule inhibitor screening in microplate-based formats. Establish baseline enzyme velocity before adding test compounds, and calculate IC₅₀ values using appropriate curve-fitting software.

3. Protein–Protein Interaction Studies: MDH2 interacts with multiple mitochondrial complex components and metabolic enzymes. The recombinant protein can be used in pull-down assays, co-immunoprecipitation controls, or surface plasmon resonance (SPR) binding studies.

4. Antibody Validation: Recombinant MDH2 protein serves as a positive control antigen for Western blotting and ELISA assay development or antibody validation workflows.

5. Structural and Biophysical Studies: The purified recombinant protein can be used for thermal shift assays (TSA/DSF), circular dichroism (CD), or as a standard for mass spectrometry-based proteomics.

Quality, Purity, and Safety Considerations

Purity: Recombinant Human MDH2 Protein from Aladdin Scientific is produced using recombinant expression systems and subjected to chromatographic purification to yield a research-grade reagent. Purity specifications are detailed on the product datasheet for SKU rp369504. SDS-PAGE analysis is typically used to confirm purity and apparent molecular weight.

Endotoxin: For cell-based or functional assays, endotoxin levels should be considered. Consult the product datasheet for endotoxin testing information. If low-endotoxin preparations are required for sensitive cell culture applications, confirm the specification prior to ordering.

Activity Verification: Where activity data are provided on the datasheet, confirm that the lot-specific certificate of analysis (CoA) is consistent with your experimental requirements before initiating assays.

Safe Handling: This product is intended for research use only and is not for diagnostic or therapeutic use in humans. Standard laboratory safety practices apply: wear appropriate PPE (gloves, lab coat, eye protection), work in a suitable environment, and dispose of materials in accordance with local regulations. The protein itself does not present a known biological hazard under normal research use conditions, but always consult the Safety Data Sheet (SDS) for complete hazard information.

Frequently asked questions

What expression system is used to produce Recombinant Human MDH2 Protein (rp369504)?

The specific expression system used for Recombinant Human MDH2 Protein (rp369504) is detailed in the product datasheet. Common expression hosts for recombinant human mitochondrial proteins include E. coli and mammalian cell systems; please consult the datasheet or contact Aladdin Scientific technical support for lot-specific information.

How should I measure MDH2 enzyme activity in vitro?

MDH2 activity is most commonly measured spectrophotometrically by monitoring the change in absorbance at 340 nm, which reflects NADH oxidation or reduction. In the standard forward (oxidative) direction, L-malate and NAD+ are used as substrates; in the reverse direction, oxaloacetate and NADH are used. Ensure substrates are freshly prepared and the assay is performed at a controlled temperature (typically 25–37 °C) with an appropriate buffer at physiological pH.

How many freeze-thaw cycles can Recombinant Human MDH2 Protein withstand without significant activity loss?

To preserve enzymatic activity, it is recommended to aliquot the reconstituted protein immediately upon first use and avoid more than 2–3 freeze-thaw cycles. Repeated freeze-thaw cycles can lead to protein aggregation and loss of activity. For extended storage, keep aliquots at −80 °C and add a stabilizing carrier protein such as 0.1% BSA if working at low concentrations.

Is Recombinant Human MDH2 Protein suitable for use as a Western blot positive control?

Yes, purified recombinant human MDH2 protein is well-suited as a positive control antigen in Western blot experiments to validate anti-MDH2 antibodies or confirm assay performance. Load an appropriate amount alongside cell lysate samples to verify antibody specificity and expected molecular weight detection.

What is the role of MDH2 in cancer metabolism research?

MDH2 has emerged as a research interest in oncology because altered TCA cycle flux and mitochondrial metabolism are hallmarks of many cancers. Studies have explored MDH2 as a potential drug target, with inhibition of MDH2 activity proposed as a strategy to disrupt cancer cell energy production. Recombinant MDH2 protein enables in vitro inhibitor screening and mechanistic studies to support this area of research.

References

1. Minarik P, Tomaskova N, Kollarova M, Antalik M. Malate dehydrogenases — structure and function. Gen Physiol Biophys. 2002;21(3):257-265. PMID:12537350
2. Wen D, Liu D, Tang J, et al. Malic enzyme 1 induces epithelial-mesenchymal transition and indicates poor prognosis in hepatocellular carcinoma. Tumour Biol. 2015;36(8):6211-6221.
3. Aladdin Scientific. Recombinant Human MDH2 Protein, SKU rp369504. Product Datasheet.

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