Cell Signaling Technology

Product Pathways - Ca / cAMP / Lipid Signaling

Phospho-PKA C (Thr197) Antibody #4781

Applications Reactivity Sensitivity MW (kDa) Source
W F E-P H M R Mk Endogenous 42 Rabbit

Applications Key:  W=Western Blotting  F=Flow Cytometry  E-P=ELISA (Peptide)
Reactivity Key:  H=Human  M=Mouse  R=Rat  Mk=Monkey
Species cross-reactivity is determined by Western blot.

Protocols

Specificity / Sensitivity

Phospho-PKA C (Thr197) Antibody detects endogenous levels of PKA C (-alpha, -beta and -gamma) only when phosphorylated at Thr197. This antibody does not cross-react with PKA C phosphorylated at other sites.

Source / Purification

Polyclonal antibodies are produced by immunizing rabbits with a synthetic phospho-peptide (KLH-coupled) corresponding to residues surrounding Thr197 of PKA C. Antibodies are purified by protein A and peptide affinity chromatography.

Western Blotting

Western Blotting

Western blot analysis of extracts from C6 and NIH/3T3 cells, untreated or treated with lambda phosphatase (PPase), using Phospho-PKA C (Thr197) Antibody (upper) or PKA C-alpha Antibody #4782 (lower).

Flow Cytometry

Flow Cytometry

Flow cytometric analysis of PC12 cells, N-ethylmaleimide (NEM) treated (blue) or Forskolin and IBMX treated (green), using Phospho-PKA C (Thr197) Antibody compared to a nonspecific negative control antibody (red).

Background

The second messenger cyclic AMP (cAMP) activates cAMP-dependent protein kinase (PKA or cAPK) in mammalian cells and controls many cellular mechanisms such as gene transcription, ion transport and protein phosphorylation (1). Inactive PKA is a heterotetramer composed of a regulatory subunit (R) dimer and a catalytic subunit (C) dimer. In this inactive state, the pseudosubstrate sequences on the R subunits block the active sites on the C subunits. Three C subunit isoforms (C-α, C-β and C-γ) and two families of regulatory subunits (RI and RII) with distinct cAMP binding properties have been identified. Within the two R families, two isoforms, α and β (RI-α, RI-β, RII-α and RII-β) exist. Upon binding of cAMP to the R subunits, the autoinhibitory contact is eased and active monomeric C subunits are released. PKA shares substrate specificity with Akt (PKB) and PKC, which is characterized by an arginine at position -3 relative to the phosphorylated serine or threonine residue (2). Substrates that present this consensus sequence and have been shown to be phosphorylated by PKA are Bad (Ser155), CREB (Ser133) and GSK-3 (GSK-3α Ser21 and GSK-3β Ser9) (3-5). In addition, combined knock-down of PKA C-α and -β blocks cAMP-mediated phosphorylation of Raf (Ser43 and Ser259) (6). Autophosphorylation and phosphorylation by PDK-1 are two known mechanisms responsible for phosphorylation of the C subunit at Thr197 (7).

  1. Montminy, M. (1997) Annu. Rev. Biochem. 66, 807-822.
  2. Dell'Acqua, M.L. and Scott, J.D. (1997) J. Biol. Chem. 272, 12881-12884.
  3. Tan, Y. et al. (2000) J. Biol. Chem. 275, 25865-25869.
  4. Gonzalez, G.A. and Montminy, M.R. (1989) Cell 59, 675-680.
  5. Fang, X. et al. (2000) Proc. Natl. Acad. Sci. USA 97, 11960-11965.
  6. Dumaz, N. and Marais, R. (2003) J. Biol. Chem. 278, 29819 -29823.
  7. Moore, M.J. et al. (2002) J. Biol. Chem. 277, 47878-47884.

Application References

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Companion Products

This product is for in vitro research use only and is not intended for use in humans or animals. This product is not intended for use as therapeutic or in diagnostic procedures.

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