A human membrane-associated folate binding protein is anchored by a glycosyl-phosphatidylinositol tail.
Membrane-associated and soluble forms of folate binding protein (FBP) have been identified in mammalian tissues and biological fluids. Despite their solubility differences, these two forms are functionally similar, immunologically cross-reacting, and have the same apparent molecular weights. In this study we demonstrate, for the first time, that the membrane FBP of cultured human KB cells contains a glycosyl-phosphatidylinositol (GPI) tail which is responsible for its hydrophobic properties and distinguishes it from the soluble FBP released into the medium. Treatment of the purified membrane FBP with phospholipase C specific for phosphatidylinositol (PI-PLC) removed the GPI tail and converted it to the soluble form without a change in apparent Mr by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In addition, virtually all of the folate binding sites on the plasma membrane of the intact cells were released as soluble, functional FBP following treatment with PI-PLC. The GPI tail contained 1-O-alkyl-2-O-acylglycerol as a mixture of fatty alcohols in ether linkage at C1 of the glycerol backbone and almost exclusively docosanoic acid (22:0) as the fatty acid on C2. The inositol also contained a mixture of fatty acids (16:0, 18:0, 18:1, 20:4, 22:0) located on a site other than the C2 position since the FBP was susceptible to PI-PLC cleavage. After nitrous acid deamination, the aqueous portion of the FBP contained covalently bound fatty acids, predominantly palmitate (16:0) and stearate (18:0), indicating the presence of additional acyl groups attached to the peptide in the form of amide, ester, or thioester linkage.
Luhrs
C A
CA
Department of Medicine, State University of New York-Health Science Center, Brooklyn.
Slomiany
B L
BL
D002352:Carrier Proteins / Q000302:isolation & purification / Q000378:metabolism*; D002462:Cell Membrane / Q000378:metabolism*; D005227:Fatty Acids / Q000032:analysis; D058973:Folate Receptors, GPI-Anchored; D005492:Folic Acid / Q000378:metabolism; D005989:Glycerides / Q000032:analysis; D006801:Humans; D007624:KB Cells; D008074:Lipoproteins / Q000302:isolation & purification / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000032:analysis*; D011956:Receptors, Cell Surface / Q000378:metabolism*; D010738:Type C Phospholipases / Q000378:metabolism
D002352:Carrier Proteins; D005227:Fatty Acids; D058973:Folate Receptors, GPI-Anchored; D005989:Glycerides; D008074:Lipoproteins; D010716:Phosphatidylinositols; D011956:Receptors, Cell Surface; D005492:Folic Acid; D010738:Type C Phospholipases