title: Bovine membranous dopamine beta-hydroxylase is not anchored via covalently attached phosphatidylinositol.
abstract: The mode of attachment of the membrane-bound form of the chromaffin granule glycoprotein dopamine beta-hydroxylase has been investigated; specifically, the possibility of a covalently attached glycosyl phosphatidylinositol anchor has been examined in detail. Incubation of fragmented chromaffin granule membranes with phosphatidylinositol-specific phospholipase C led to no solubilization of dopamine beta-hydroxylase activity. Further, chemical analyses of the inositol and phosphate content of the soluble form of dopamine beta-hydroxylase indicate that the enzyme contains no covalently attached phosphate or inositol. The results of these studies allow elimination of phosphatidylinositol as a source of membrane anchoring for dopamine beta-hydroxylase and, coupled with previously available data, argue against a post-translational covalent modification of the enzyme as the mechanism of membrane attachment. Possible alternative models for the anchoring of membranous dopamine beta-hydroxylase are discussed.
authors.affiliation: Department of Chemistry, University of California, Berkeley 94720.
authors.forename: L C
authors.initials: LC
authors.lastname: Stewart
authors.forename: J P
authors.initials: JP
authors.lastname: Klinman
mesh_terms: D000818:Animals; D002417:Cattle; D002837:Chromaffin Granules / Q000201:enzymology*; D002838:Chromaffin System / Q000201:enzymology*; D004299:Dopamine beta-Hydroxylase / Q000032:analysis / Q000378:metabolism*; D008401:Gas Chromatography-Mass Spectrometry; D007294:Inositol / Q000032:analysis; D007425:Intracellular Membranes / Q000201:enzymology; D010710:Phosphates / Q000032:analysis; D010716:Phosphatidylinositols / Q000378:metabolism*; D011499:Protein Processing, Post-Translational; D012995:Solubility; D013211:Staphylococcus aureus / Q000201:enzymology; D010738:Type C Phospholipases / Q000378:metabolism
chemical_list: D010710:Phosphates; D010716:Phosphatidylinositols; D007294:Inositol; D004299:Dopamine beta-Hydroxylase; D010738:Type C Phospholipases