Two membrane-bound forms of tyrosylprotein sulfotransferase as revealed by phase partitioning in Triton X-114.
Tyrosylprotein sulfotransferase (TPST) is a membrane-associated enzyme of the trans Golgi network that catalyzes the posttranslational sulfation of a variety of secretory and membrane proteins. We have analyzed the membrane association of TPST in Golgi-enriched fractions from bovine adrenal medulla using carbonate treatment (pH 11) and Triton X-114 phase partitioning. TPST was not extracted by carbonate. Triton X-114 phase partitioning revealed that, unexpectedly, TPST from non-carbonate-treated membranes was present in both, a hydrophilic and a hydrophobic form with apparent sedimentation coefficients of approximately 13 and approximately 6, respectively. Extraction of membranes with carbonate converted the hydrophilic form TPST to the hydrophobic form. Addition of the carbonate extract to TPST solubilized from carbonate-treated membranes converted the hydrophobic form of the enzyme to the hydrophilic form. This conversion of TPST was specific in that it was not observed for the bulk of the proteins present in the carbonate-treated membranes. The factor in the carbonate extract responsible for this conversion, referred to as "phase-transfer factor", (i) was precipitable with ammonium sulfate and polyethylene glycol, (ii) was non-dialyzable, (iii) was not extracted from membranes by 0.5 M NaCl, and (iv) appeared to be more abundant than TPST itself. These results show that TPST is an integral membrane protein and suggested that the enzyme may exist in a complex with a peripheral membrane protein. Moreover, a phase-transfer factor was also observed in another system, PC12 cells.(ABSTRACT TRUNCATED AT 250 WORDS)
Niehrs
C
C
Cell Biology Programme, European Molecular Biology Laboratory, Heidelberg/Germany.
Stinchcombe
J C
JC
Huttner
W B
WB
D000313:Adrenal Medulla / Q000201:enzymology; D000818:Animals; D002417:Cattle; D002499:Centrifugation, Density Gradient; D003902:Detergents; D006056:Golgi Apparatus / Q000201:enzymology*; D046911:Macromolecular Substances; D017830:Octoxynol; D011092:Polyethylene Glycols; D015238:Sulfotransferases / Q000302:isolation & purification*
D003902:Detergents; D046911:Macromolecular Substances; D011092:Polyethylene Glycols; D017830:Octoxynol; C010615:Nonidet P-40; D015238:Sulfotransferases; C039079:protein-tyrosine sulfotransferase