Phospholipid is required for the processing of presecretory proteins by detergent-solubilized canine pancreatic signal peptidase.
The ability of canine pancreatic signal peptidase to remove the signal peptide portion of presecretory proteins in a translocation-independent assay is shown to require phospholipid. Sodium deoxycholate extracts of canine pancreatic rough microsomes containing both signal peptidase and phospholipid were delipidated by gel filtration chromatography on Sepharose CL-6B equilibrated with 0.2% deoxycholate. Column fractions were assayed for signal peptidase activity both with and without the addition of ethanol-extracted soybean phospholipid at a final concentration of 1.0 mg/ml. A peak of signal peptidase activity was detected only when the fractions were assayed with added phospholipid. Phospholipid assays demonstrated that the peak of signal peptidase activity was cleanly separated from phospholipid. The ratio of protein to phospholipid in the deoxycholate extract of rough microsomes was 1.76 while that of the most active signal peptidase fractions ranged from 46.1 to 138. The peak of signal peptidase activity exhibited an apparent Stokes radius of 55 A. Highly purified preparations of phosphatidylcholine were most effective in restoring activity to delipidated signal peptidase. Phosphatidylinositol was much less effective. Phosphatidylserine, phosphatidylethanolamine, sphingomyelin, and lysophosphatidylcholine were all ineffective.
D000818:Animals; D002417:Cattle; D003840:Deoxycholic Acid / Q000302:isolation & purification; D003902:Detergents; D004285:Dogs; D010450:Endopeptidases / Q000378:metabolism*; D008081:Liposomes; D008565:Membrane Proteins; D008861:Microsomes / Q000201:enzymology; D010179:Pancreas / Q000201:enzymology*; D010743:Phospholipids / Q000494:pharmacology*; D010944:Plants / Q000378:metabolism; D014176:Protein Biosynthesis; D011498:Protein Precursors / Q000096:biosynthesis*; D012697:Serine Endopeptidases; D013025:Glycine max; D014908:Triticum / Q000378:metabolism
D003902:Detergents; D008081:Liposomes; D008565:Membrane Proteins; D010743:Phospholipids; D011498:Protein Precursors; D003840:Deoxycholic Acid; D010450:Endopeptidases; D012697:Serine Endopeptidases; C026515:type I signal peptidase