Abstract |
A major glycoprotein of rat hepatoma plasma membranes was selectively released as a soluble form by incubating the membrane with phosphatidylinositol-specific phospholipase C. The soluble form corresponding to the glycoprotein was also prepared by butan-1-ol extraction of microsomal membranes at pH 5.5, whereas extraction at pH 8.5 yielded an electrophoretically different form with a hydrophobic nature. The soluble glycoprotein extracted at pH 5.5 was purified by sequential chromatography on concanavalin A-Sepharose, Sephacryl S-300 and anti-( alkaline phosphatase) IgG-Sepharose, the last step being used to remove a contaminating alkaline phosphatase. The glycoprotein thus purified was a single protein with Mr 130,000 in SDS/ polyacrylamide-gel electrophoresis, although it behaved as a dimer in gel filtration on Sephacryl S-300. The glycoprotein was analysed for amino acid and carbohydrate composition. The composition of the carbohydrate moiety, which amounted to 64% by weight, suggested that the glycoprotein contained much larger numbers of N-linked oligosaccharide chains than those with O-linkage. It was confirmed that the purified glycoprotein was immunologically identical not only with that released by the phospholipase C but also with the hydrophobic form extracted with butan-1-ol at pH 8.5. The results indicate that the glycoprotein of rat hepatoma plasma membranes, which has an unusually high content of carbohydrate, is another membrane protein released by phosphatidylinositol-specific phospholipase C, as documented for alkaline phosphatase, acetylcholinesterase and Thy-1 antigen.
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Authors | Y Ikehara, Y Hayashi, S Ogata, A Miki, T Kominami |
Journal | The Biochemical journal
(Biochem J)
Vol. 241
Issue 1
Pg. 63-70
(Jan 01 1987)
ISSN: 0264-6021 [Print] England |
PMID | 3032162
(Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
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Chemical References |
- Amino Acids
- Butanols
- Carbohydrates
- Glycoproteins
- Membrane Glycoproteins
- Membrane Proteins
- Neoplasm Proteins
- hepatoma adhesive factor, rat
- 1-Butanol
- Alkaline Phosphatase
- Phosphoric Diester Hydrolases
- Phosphoinositide Phospholipase C
- Phosphatidylinositol Diacylglycerol-Lyase
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Topics |
- 1-Butanol
- Alkaline Phosphatase
(metabolism)
- Amino Acids
(analysis)
- Animals
- Butanols
(pharmacology)
- Carbohydrates
(analysis)
- Cell Line
- Cell Membrane
(drug effects, enzymology)
- Chromatography, Gel
- Electrophoresis, Polyacrylamide Gel
- Glycoproteins
(isolation & purification)
- Membrane Glycoproteins
- Membrane Proteins
(isolation & purification)
- Neoplasm Proteins
(isolation & purification)
- Phosphatidylinositol Diacylglycerol-Lyase
- Phosphoinositide Phospholipase C
- Phosphoric Diester Hydrolases
(pharmacology)
- Rats
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