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Active Sphingosine Kinase 2 (SPHK2)

ACTIVITY TEST

Sphingosine Kinase 2 (SPHK2) is an enzyme that phosphorylates sphingosine to produce sphingosine-1-phosphate (S1P), a bioactive lipid mediator involved in cell proliferation, survival, and immune regulation. Unlike its isoform SPHK1, which is primarily cytosolic, SPHK2 localizes to the nucleus, mitochondria, and endoplasmic reticulum, suggesting distinct roles in cellular processes. SPHK2 regulates key pathways such as apoptosis, autophagy, and epigenetic modulation by influencing histone acetylation. It has been implicated in diseases like cancer, neurodegeneration, and metabolic disorders, where its activity can either promote or suppress pathology depending on context.Besides,Sphingosine 1 Phosphate Lyase 1 (SGPL1) has been identified as an interactor of SPHK2, thus a functional binding ELISA assay was conducted to detect the interaction of recombinant human SPHK2 and recombinant human SGPL1. Briefly, SPHK2 was diluted serially in PBS with 0.01% BSA (pH 7.4). Duplicate samples of 100 μl were then transferred to SGPL1-coated microtiter wells and incubated for 1h at 37℃. Wells were washed with PBST and incubated for 1h with anti-SPHK2 pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody for 1h at 37℃, wells were aspirated and washed 5 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50 µL stop solution to the wells and read at 450/630nm immediately. The binding activity of recombinant human SPHK2 and recombinant human SGPL1 was shown in Figure 1, the EC50 for this effect is 0.08µg/mL.

USAGE

Reconstitute in 10mM PBS (pH7.4) to a concentration of 0.1-1.0 mg/mL. Do not vortex.

STORAGE

Avoid repeated freeze/thaw cycles. Store at 2-8°C for one month. Aliquot and store at -80°C for 12 months.

STABILITY

The thermal stability is described by the loss rate. The loss rate was determined by accelerated thermal degradation test, that is, incubate the protein at 37°C for 48h, and no obvious degradation and precipitation were observed. The loss rate is less than 5% within the expiration date under appropriate storage condition.

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