Phospholipase B from the plasma membrane of Saccharomyces cerevisiae. Separation of two forms with different carbohydrate content.
Two forms of phospholipase B could be solubilized from the plasma membrane of Saccharomyces cerevisiae, separated by gel filtration with Sephacryl S-300 and identified by SDS-polyacrylamide gel electrophoresis as glycoproteins of the apparent molecular weights of about 220 000 (phospholipase B1) and 145 000 (phospholipase B2). The enzymes are very similar in respect to their catalytic properties. Both forms converted lysophosphatidylcholine to diacylphosphatidylcholine and unesterified fatty acids. The carbohydrate content of the glycoproteins could be reduced by treatment with endoglycosidase H and HF. By incubation of phospholipase B1 and phospholipase B2 with endoglycosidase H from Streptomyces griseus, one main protein with an apparent Mr of 67 000 and the same residual carbohydrate content was obtained. Treatment with HF reduced phospholipase B1 and phospholipase B2 to proteins with an apparent Mr of 52 000 and 67 000, respectively. These results could indicate that the two forms are similar in respect to their protein moieties. An antiserum raised in mice against phospholipase B2 showed no crossreactivity with phospholipase B1 as detected by immunoblot analysis. The reactivity of phospholipase B2 was diminished or abolished by progressive removal of carbohydrate. These results were taken as indications for differences in the carbohydrate component of the two enzyme forms.
D002462:Cell Membrane / Q000201:enzymology; D006023:Glycoproteins / Q000302:isolation & purification; D008245:Lysophospholipase / Q000276:immunology / Q000302:isolation & purification*; D008970:Molecular Weight; D010740:Phospholipases / Q000302:isolation & purification*; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D006023:Glycoproteins; D010740:Phospholipases; D008245:Lysophospholipase