1. Academic Validation
  2. A phosphatase resistant substrate for the assay of protein kinase C in crude tissue extracts

A phosphatase resistant substrate for the assay of protein kinase C in crude tissue extracts

  • Biochem Biophys Res Commun. 1991 Oct 31;180(2):694-701. doi: 10.1016/s0006-291x(05)81121-0.
Y J Farrar 1 T C Vanaman J T Slevin
Affiliations

Affiliation

  • 1 Veterans Administration Medical Center, Lexington, KY 40511.
Abstract

Protein kinase C (PKC) is routinely assayed, after it is partially purified over DEAE-cellulose chromatography to eliminate any interfering protein kinases and phosphatases, by measuring the transfer of gamma-phosphate of [gamma-32P]ATP to H1 histone. Recently, it has been shown that a synthetic peptide, comprising residues 4-14 of myelin basic protein (MBP4-14), is a very selective PKC substrate which is not phosphorylated effectively by cyclic AMP-dependent protein kinase, Casein Kinase I and II, Ca2+/Calmodulin dependent protein kinase II or phosphorylase kinase [Yasuda, I., Kishimoto, A., Tanaka, S-I., Tominaga, M., Sakurai, A. and Nishizuka, Y. (1990) BBRC 166, 1220-1227]. We report here that once MBP4-14 is phosphorylated, it is not dephosphorylated by okadaic acid-sensitive phosphatases (protein phosphatases 1, 2A and 3) or other protein phosphatases such as Calcineurin and/or PP 2C present in hippocampal homogenates. Therefore, MBP4-14 can be used for PKC assay in crude extracts of neural tissue.

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