ADP-Glo Kinase Assay + PDK1 Kinase Enzyme System
Recombinant full-length human PDK1 was expressed by baculovirus in Sf9 insect cells using an N-terminal His tag. PDK1 (3-phosphoinositide-dependent protein kinase) is activated by the presence of PtdIns(3,4,5)P3 or PtdIns(3,4)P2. PDK1 then activates protein kinase B (PKB), which in turn, inactivates glycogen synthase kinase-3 (GSK3). The phosphorylation of other proteins by PKB and GSK3 is likely to mediate many of the intracellular actions of insulin. Thus, PDK1 plays a key role in mediating many of the actions of the second messenger(s) PtdIns(3,4,5)P3 and/or PtdIns(3,4)P2. The human PDK1 is a 556-residue monomeric enzyme comprising of a catalytic domain that is most similar to the PKA, PKB and PKC subfamily of protein kinases. 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: PDK1, 10ug (Human, recombinant full-length). MW: ~67kDa. Substrate: PDKtide (KTFCGTPEYLAPEVRREPRILSEEEQEMFRDFDYIADWC); derived from two human proteins: residues 1-14 are based on AKT1 (307-320) and residues 16-39 are based on PKN2/PRK2 (961-984). Other: Reaction Buffer, DTT. PDK1 NCBI Database Entry: www.ncbi.nlm.nih.gov/gene/5170/. Visit www.promega.com/kinase/ to see all Kinase Enzyme Systems.