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.