Reconstitution of Saccharomyces cerevisiae phosphatidylserine synthase into phospholipid vesicles. Modulation of activity by phospholipids.
Membrane-associated phosphatidylserine synthase was purified from Saccharomyces cerevisiae (Bae-Lee, M., and Carman, G. M. (1984) J. Biol. Chem. 259, 10857-10862) and reconstituted into phospholipid vesicles containing phosphatidylcholine/phosphatidylethanolamine/ phosphatidylinositol/phosphatidylserine. Reconstitution was performed by removing detergent from an octyl glucoside/phospholipid/Triton X-100/enzyme mixed micelle by Sephadex G-50 super-fine chromatography. The average diameter of the vesicles was 90 nm, and the enzyme was reconstituted asymmetrically with the active site facing outward. The enzymological properties of reconstituted phosphatidylserine synthase were determined in the absence of detergent. The enzyme was reconstituted into vesicles with phospholipid compositions approximating those of wild type and mutant strains of S. cerevisiae. Reconstituted activity was modulated by the phosphatidylinositol/phosphatidylserine ratio in the vesicles. The modulation of activity observed in the vesicles is enough to account for some of the fluctuations in the phosphatidylserine content in vivo.
D001665:Binding Sites; D010717:CDPdiacylglycerol-Serine O-Phosphatidyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002850:Chromatography, Gel; D002918:Chymotrypsin / Q000494:pharmacology; D005960:Glucosides; D008081:Liposomes / Q000378:metabolism*; D008823:Micelles; D017830:Octoxynol; D010713:Phosphatidylcholines / Q000494:pharmacology; D010714:Phosphatidylethanolamines / Q000494:pharmacology; D010716:Phosphatidylinositols / Q000494:pharmacology; D010718:Phosphatidylserines / Q000494:pharmacology; D010743:Phospholipids / Q000494:pharmacology*; D010770:Phosphotransferases / Q000378:metabolism*; D011092:Polyethylene Glycols; D012441:Saccharomyces cerevisiae / Q000201:enzymology*
D005960:Glucosides; D008081:Liposomes; D008823:Micelles; D010713:Phosphatidylcholines; D010714:Phosphatidylethanolamines; D010716:Phosphatidylinositols; D010718:Phosphatidylserines; D010743:Phospholipids; C018619:octyl-beta-D-glucoside; D011092:Polyethylene Glycols; D017830:Octoxynol; D010770:Phosphotransferases; D010717:CDPdiacylglycerol-Serine O-Phosphatidyltransferase; D002918:Chymotrypsin