Phosphatidate phosphatase from Saccharomyces cerevisiae. Isolation of 45- and 104-kDa forms of the enzyme that are differentially regulated by inositol.
Immunoblot analysis of cell extracts using antibodies specific for the 91-kDa form of membrane-associated phosphatidate phosphatase from Saccharomyces cerevisiae (Lin, Y.-P., and Carman, G.M. (1989) J. Biol. Chem. 264, 8641-8645) revealed the existence of a 45-kDa form of the enzyme. Immunoblot analysis also showed that the 91-kDa form of the enzyme was a proteolytic product of a 104-kDa enzyme. The mitochondrial fraction contained the 45-kDa enzyme, whereas the microsomal fraction contained the 45- and 104-kDa enzymes. In vivo labeling experiments showed that the 104-kDa form of phosphatidate phosphatase was not a precursor of the 45-kDa form of the enzyme. The 45- and 104-kDa forms of phosphatidate phosphatase were purified and characterized. The enzymological properties of both enzymes were similar. However, the phosphatidate phosphatase 45- and 104-kDa proteins differed with respect to their isoelectric points and peptide fragments resulting from V8 proteolysis and cyanogen bromide cleavage. The expression of the phosphatidate phosphatase 45- and 104-kDa enzymes were regulated differentially in cells supplemented with inositol. The addition of inositol to the growth medium resulted in the induction of the phosphatidate phosphatase 45-kDa enzyme. The expression of the 104-kDa enzyme was not affected by inositol. Both forms of phosphatidate phosphatase were induced when cells entered the stationary phase of growth.
Morlock
K R
KR
Department of Food Science, Cook College, Rutgers University, New Brunswick 08903.
McLaughlin
J J
JJ
Lin
Y P
YP
Carman
G M
GM
D015153:Blotting, Western; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007294:Inositol / Q000378:metabolism; D007525:Isoelectric Focusing; D007527:Isoenzymes / Q000302:isolation & purification*; D008274:Magnesium; D017830:Octoxynol; D010658:Phenylglyoxal / Q000494:pharmacology; D010711:Phosphatidate Phosphatase / Q000037:antagonists & inhibitors / Q000302:isolation & purification* / Q000378:metabolism; D011092:Polyethylene Glycols; D011233:Precipitin Tests; D011433:Propranolol / Q000494:pharmacology; D012441:Saccharomyces cerevisiae / Q000201:enzymology*; D013696:Temperature
D003902:Detergents; D007527:Isoenzymes; D011092:Polyethylene Glycols; D007294:Inositol; D017830:Octoxynol; D011433:Propranolol; D010711:Phosphatidate Phosphatase; D008274:Magnesium; D010658:Phenylglyoxal