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Active Growth Arrest And DNA Damage Inducible Protein Alpha (GADD45a)

DDIT1; GADD45; DNA damage-inducible transcript 1 protein

ACTIVITY TEST

Growth arrest and DNA-damage-inducible protein GADD45 alpha (GADD45a) is a member of the GADD45 family. It may affect PCNA interaction with some CDK (cell division protein kinase) complexes; stimulates DNA excision repair in vitro and inhibits entry of cells into S phase. In T-cells, functions as a regulator of p38 MAPKs by inhibiting p88 phosphorylation and activity. Besides, Proliferating Cell Nuclear Antigen (PCNA) has been identified as an interactor of GADD45a, thus a binding ELISA assay was conducted to detect the interaction of recombinant rat GADD45a and recombinant rat PCNA. Briefly, GADD45a were diluted serially in PBS, with 0.01% BSA (pH 7.4). Duplicate samples of 100μL were then transferred to PCNA-coated microtiter wells and incubated for 2h at 37℃. Wells were washed with PBST and incubated for 1h with anti-GADD45a pAb, then aspirated and washed 3 times. After incubation with HRP labelled secondary antibody, wells were aspirated and washed 3 times. With the addition of substrate solution, wells were incubated 15-25 minutes at 37℃. Finally, add 50µL stop solution to the wells and read at 450nm immediately. The binding activity of GADD45a and PCNA was shown in Figure 1, and this effect was in a dose dependent manner.
Figure. The binding activity of GADD45a with PCNA.

USAGE

Reconstitute in 20mM Tris, 150mM NaCl (pH8.0) 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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