PURIFICATION OF THE GOLGI ADENOSINE 3'-PHOSPHATE 5'-PHOSPHOSULFATE TRANSPORTER, A HOMODIMER WITHIN THE MEMBRANE

Sulfation of proteoglycans, secretory and membrane proteins, and glyco lipids occurs in the lumen of the Golgi apparatus. Adenosine 3'-phosph ate 5'-phosphosulfate (PAPS), the sulfate donor in these reactions, mu st be transported from the cytosol, its site of synthesis, into the lu men of the Golgi apparatus. We have identified and purified to apparen t homogeneity the rat liver Golgi membrane PAPS transporter by a combi nation of conventional and affinity chromatography as well as photoaff inity radiolabeling with adenosine 3',5'-bisphosphate, a competitive i nhibitor of PAPS transport. The transporter, a 75-kDa protein, was pur ified 70,000-fold over homogenate (6% yield) and transported PAPS into phosphatidylcholine liposomes selectively and in a saturable manner ( apparent K-m of 1.7 mu M). Radiation target inactivation analyses of t he transport activity in rat liver Golgi vesicles, together with the a bove described biochemical apl preaches, demonstrate that the PAPS tra nsporter within the Golgi membrane is a homodimer.