THE DETECTION OF PHOSPHOLIPASE-RESISTANT AND PHOSPHOLIPASE-SENSITIVE GLYCOSYL-PHOSPHATIDYLINOSITOL MEMBRANE ANCHORS BY WESTERN BLOTTING

Glycosyl-phosphatidylinositol (GPI) membrane anchors are present on a large number of eukaryotic plasma membrane proteins. Some of these anc hors can be cleaved with bacterial phosphatidylinositol-specific phosp holipases C, and a glycosyl-phosphatidylinositol-specific phospholipas e C from Trypanosoma brucei, to reveal an epitope called the cross-rea cting determinant. Other glycosyl-phosphatidylinositol anchors are res istant to the action of these enzymes prior to treatment with mild bas e. A simple method is described for identifying both phospholipase-sen sitive and -resistant anchors using anti-cross-reacting determinant an tibodies on Western blots. This procedure represents a high-sensitivit y general method for the identification of GPI-anchored proteins. (C) 1994 Academic Press, Inc.