Attachment of human placental-type alkaline phosphatase via phosphatidylinositol to syncytiotrophoblast and tumour cell plasma membranes.
Phosphatidylinositol anchors human placental-type alkaline phosphatase (PLAP) to both syncytiotrophoblast and tumour cell plasma membranes. PLAP activity was released from isolated human placental syncytiotrophoblast plasma membranes and the surface of tumour cells with a phospholipase C from Bacillus cereus. This was a specific event, not the result of proteolysis or membrane perturbation, but the action of a phosphatidylinositol-specific phospholipase C in the preparation. Soluble PLAP, released with B. cereus phospholipase C and purified by immunoaffinity chromatography, ran on SDS-PAGE as a 66-kDa band. This corresponded to intact PLAP molecules. The protease bromelain cleaved lower-molecular-mass PLAP (64 kDa) from the membranes. Flow cytometry demonstrated that B. cereus phospholipase C released human tumour cell membrane PLAP in preference to other cell-surface molecules. This was in contrast to the non-specific proteolytic action of bromelain or Clostridium perfringens phospholipase C, which had no effect on membrane PLAP expression. Radiolabelling of tumour cells with fatty acids indicated PLAP to be labelled with both [3H]myristic and [3H]palmitic acid. This fatty-acid--PLAP bond was sensitive to pH 10 hydroxylamine treatment indicating an O-ester linkage.
D000469:Alkaline Phosphatase / Q000378:metabolism*; D001409:Bacillus cereus / Q000201:enzymology; D001963:Bromelains / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D003013:Clostridium / Q000201:enzymology; D005260:Female; D058851:GPI-Linked Proteins; D006801:Humans; D007527:Isoenzymes / Q000378:metabolism*; D010716:Phosphatidylinositols / Q000494:pharmacology*; D011247:Pregnancy; D011990:Receptors, Transferrin / Q000378:metabolism; D014327:Trophoblasts / Q000378:metabolism*; D014407:Tumor Cells, Cultured; D010738:Type C Phospholipases / Q000494:pharmacology
D058851:GPI-Linked Proteins; D007527:Isoenzymes; D010716:Phosphatidylinositols; D011990:Receptors, Transferrin; D001963:Bromelains; D000469:Alkaline Phosphatase; C019839:alkaline phosphatase, placental; D010738:Type C Phospholipases