Phosphatidylinositol anchor of HeLa cell alkaline phosphatase

Journal Article · Tue Sep 08 00:00:00 EDT 1987 · Biochemistry; (United States)

DOI:https://doi.org/10.1021/bi00392a019· OSTI ID:5721146

Jemmerson, R; Low, M G

Alkaline phosphatase from cancer cells, HeLa TCRC-1, was biosynthetically labeled with either /sup 3/H-fatty acids or (/sup 3/H)ethanolamine as analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and fluorography of immunoprecipitated material. Phosphatidylinositol-specific phospholipase C (PI-PLC) released a substantial proportion of the /sup 3/H-fatty acid label from immunoaffinity-purified alkaline phosphatase but had no effect on the radioactivity of (/sup 3/H)ethanolamine-labeled material. PI-PLC also liberated catalytically active alkaline phosphatase from viable cells, and this could be selectively blocked by monoclonal antibodies to alkaline phosphatase. However, the alkaline phosphatase released from /sup 3/H-fatty acid labeled cells by PI-PLC was not radioactive. By contrast, treatment with bromelain removed both the /sup 3/H-fatty acid and the (/sup 3/H)ethanolamine label from purified alkaline phosphatase. Subtilisin was also able to remove the (/sup 3/H)ethanolamine label from the purified alkaline phosphatase. The /sup 3/H radioactivity in alkaline phosphatase purified from (/sup 3/H)ethanolamine-labeled cells comigrated with authentic (/sup 3/H)ethanolamine by anion-exchange chromatography after acid hydrolysis. The data suggest that the /sup 3/H-fatty acid and (/sup 3/H)ethanolamine are covalently attached to the carboxyl-terminal segment since bromelain and subtilisin both release alkaline phosphatase from the membrane by cleavage at that end of the polypeptide chain. The data are consistent with findings for other proteins recently shown to be anchored in the membrane through a glycosylphosphatidylinositol structure and indicate that a similar structure contributes to the membrane anchoring of alkaline phosphatase.

• Research Organization: Univ. of Minnesota, Minneapolis
• OSTI ID: 5721146
• Journal Information: Biochemistry; (United States), Vol. 26:18
• Country of Publication: United States
• Language: English

Related Subjects

62 RADIOLOGY AND NUCLEAR MEDICINE
ALKALINE PHOSPHATASE
AUTORADIOGRAPHY
BIOSYNTHESIS
TRITIUM COMPOUNDS
AMINES
CARBOXYLIC ACIDS
ELECTROPHORESIS
HELA CELLS
TUMOR CELLS
ANIMAL CELLS
ENZYMES
ESTERASES
HYDROLASES
LABELLED COMPOUNDS
ORGANIC ACIDS
ORGANIC COMPOUNDS
PHOSPHATASES
SYNTHESIS
550601* - Medicine- Unsealed Radionuclides in Diagnostics