Purification and characterization of ceramide-activated protein phosphatases

Sehamuddin Galadari, Katsuya Kishikawa, Craig Kamibayashi, Marc C. Mumby, Yusuf A. Hannun

Research output: Contribution to journalArticle

Abstract

Ceramide has emerged as a potential regulator of diverse cellular functions, and a few direct targets have been identified for its action including protein kinases and phosphatases. In this study, we have purified the predominant ceramide-activated protein phosphatase (CAPP) from rat brain. Utilizing a novel chromatographic approach, CAPP was purified to near homogeneity using hydrophobic interaction chromatography on phenyl Sepharose followed by anion-exchange chromatography on MonoQ. The purified protein was composed of three major bands on SDS-polyacrylamide gel electrophoresis which comigrated with the three subunits of heterotrimeric PP2A. Immunologic studies further identified CAPP to be composed predominantly of heterotrimeric AB'C and ABαC as well as heterodimeric PP2A (AC), where C is the catalytic subunit, and A and B are regulatory subunits. These results were also supported by the coelution of CAPP with trimeric and dimeric PP2A on size-exclusion chromatography. These studies provide a convenient and efficient method for the isolation of trimeric and dimeric PP2A, and they allow the biochemical investigation of CAPP.

Original languageEnglish (US)
Pages (from-to)11232-11238
Number of pages7
JournalBiochemistry
Volume37
Issue number32
DOIs
StatePublished - Aug 11 1998

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Ceramides
Phosphoprotein Phosphatases
Purification
Chromatography
Size exclusion chromatography
Electrophoresis
Hydrophobic and Hydrophilic Interactions
Protein Kinases
Gel Chromatography
Anions
Rats
Polyacrylamide Gel Electrophoresis
Brain
Catalytic Domain
Proteins

ASJC Scopus subject areas

  • Biochemistry

Cite this

Galadari, S., Kishikawa, K., Kamibayashi, C., Mumby, M. C., & Hannun, Y. A. (1998). Purification and characterization of ceramide-activated protein phosphatases. Biochemistry, 37(32), 11232-11238. https://doi.org/10.1021/bi980911+

Purification and characterization of ceramide-activated protein phosphatases. / Galadari, Sehamuddin; Kishikawa, Katsuya; Kamibayashi, Craig; Mumby, Marc C.; Hannun, Yusuf A.

In: Biochemistry, Vol. 37, No. 32, 11.08.1998, p. 11232-11238.

Research output: Contribution to journalArticle

Galadari, S, Kishikawa, K, Kamibayashi, C, Mumby, MC & Hannun, YA 1998, 'Purification and characterization of ceramide-activated protein phosphatases', Biochemistry, vol. 37, no. 32, pp. 11232-11238. https://doi.org/10.1021/bi980911+
Galadari, Sehamuddin ; Kishikawa, Katsuya ; Kamibayashi, Craig ; Mumby, Marc C. ; Hannun, Yusuf A. / Purification and characterization of ceramide-activated protein phosphatases. In: Biochemistry. 1998 ; Vol. 37, No. 32. pp. 11232-11238.
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