Separation of endocytic from exocytic coated vesicles using a novel cholinesterase mediated density shift technique.
Coated vesicles isolated from rat liver perfused with diisopropylfluorophosphate (DFP) to inactivate endogenous cholinesterase contained newly synthesized secretory cholinesterase after a 30 min recovery. The cholinesterase is found in coated vesicles of presumed endocytic origin following DFP treatment and perfusion for 3 min with galactosylated cholinesterase, a ligand for the asialoglycoprotein receptor. Highly enriched populations of endocytic and exocytic coated vesicles can be separated by use of a novel cholinesterase mediated density shift technique. The two coated vesicle classes have very similar polypeptide compositions but differ significantly in the ratio of cholesterol to phospholipid.
Helmy
S
S
Porter-Jordan
K
K
Dawidowicz
E A
EA
Pilch
P
P
Schwartz
A L
AL
Fine
R E
RE
D000110:Acetylcholinesterase / Q000378:metabolism; D000818:Animals; D002091:Butyrylcholinesterase / Q000378:metabolism; D002458:Cell Fractionation; D002499:Centrifugation, Density Gradient; D002784:Cholesterol / Q000032:analysis; D004591:Electrophoresis, Polyacrylamide Gel; D004705:Endocytosis; D011992:Endosomes / Q000032:analysis / Q000201:enzymology / Q000378:metabolism*; D005089:Exocytosis; D005690:Galactose / Q000378:metabolism; D007531:Isoflurophate / Q000494:pharmacology; D008099:Liver / Q000648:ultrastructure*; D010743:Phospholipids / Q000032:analysis; D051381:Rats; D011919:Rats, Inbred Strains
D010743:Phospholipids; D007531:Isoflurophate; D002784:Cholesterol; D000110:Acetylcholinesterase; D002091:Butyrylcholinesterase; D005690:Galactose