Purification of the Golgi adenosine 3'-phosphate 5'-phosphosulfate transporter, a homodimer within the membrane
UMass Chan AffiliationsDepartment of Biochemistry and Molecular Pharmacology
Document TypeJournal Article
Chromatography, High Pressure Liquid
Chromatography, Ion Exchange
Dose-Response Relationship, Radiation
Electrophoresis, Polyacrylamide Gel
Medicine and Health Sciences
MetadataShow full item record
AbstractSulfation of proteoglycans, secretory and membrane proteins, and glycolipids occurs in the lumen of the Golgi apparatus. Adenosine 3'-phosphate 5'-phosphosulfate (PAPS), the sulfate donor in these reactions, must be transported from the cytosol, its site of synthesis, into the lumen of the Golgi apparatus. We have identified and purified to apparent homogeneity the rat liver Golgi membrane PAPS transporter by a combination of conventional and affinity chromatography as well as photoaffinity radiolabeling with adenosine 3',5'-bisphosphate, a competitive inhibitor of PAPS transport. The transporter, a 75-kDa protein, was purified 70,000-fold over homogenate (6% yield) and transported PAPS into phosphatidylcholine liposomes selectively and in a saturable manner (apparent Km of 1.7 microM). Radiation target-inactivation analyses of the transport activity in rat liver Golgi vesicles, together with the above described biochemical approaches, demonstrate that the PAPS transporter within the Golgi membrane is a homodimer.
Proc Natl Acad Sci U S A. 1994 Oct 25;91(22):10707-11.