Recombinant full-length human NEK3 was expressed by baculovirus in Sf9 insect cells using an N-terminal GST tag. NEK3 is a member of the NEK family of protein kinases that share high amino acid homology with NIMA (never in mitosis gene a). NEK3 mRNA is detected in all the proliferating cell lines with the amount not changing during the cell cycle (1). During Prolactin receptor signaling, VAV2 is phosphorylated and activated by NEK3. Overexpression of NEK3 in Chinese hamster ovary cells increases cytoskeletal re-organization in response to Prolactin while downregulation of NEK3 expression by siRNA blocks these effects (2). Prolactin also stimulates interaction between NEK3 and paxillin leading to increased paxillin phosphorylation. Analysis of breast tissue microarrays show a significant up-regulation of NEK3 expression in malignant versus normal specimens.
ADP-Glo™ Kinase Assay is a luminescent kinase assay that measures ADP formed from a kinase reaction; ADP is converted into ATP, which is a substrate in a reaction catalyzed by Ultra-Glo™ Luciferase that produces light. The luminescent signal positively correlates with ADP amount and kinase activity. The assay is well suited for measuring the effects chemical compounds have on the activity of a broad range of purified kinases, making it ideal for both primary screening as well as kinase selectivity profiling. The ADP-Glo™ Kinase Assay can be used to monitor the activity of virtually any ADP-generating enzyme (e.g., kinase or ATPase) using up to 1mM ATP.
Kinase Enzyme System contains:
Kinase: NEK3, 10μg (Human,recombinant full length). MW: ~86kDa. Substrate: Native Swine Myelin Basic Protein (MBP). Other: Reaction Buffer, DTT.
Profile More Compounds In-House: ADP-Glo™ Kinase Assay + Kinase Enzyme System is optimized so that you are up and running in no time. Complete Systems: The Kinase Enzyme Systems include a recombinant kinase enzyme, a substrate appropriate for the enzyme, a reaction buffer, DTT and supplemental reagents as needed. Obtain Reliable Results: The broad dynamic range, the ease of use and better sensitivity obtained with ADP-Glo™ Kinase Assay result in less ambiguous data.
Notes
Kinase Enzyme System manufactured by SignalChem.
Bulk quantities available upon request.
References
1.Kimura, M. et al. (2001) Molecular cloning and characterization of the human NIMA-related protein kinase 3 gene (NEK3). Cytogenet Cell Genet. 95, 177–82.
2.Miller, S.L. et al. (2007) Nek3 kinase regulates prolactin-mediated cytoskeletal reorganization and motility of breast cancer cells. Oncogene 26, 4668–78.
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