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EMBO J. Jan 1990; 9(1): 35–42.
PMCID: PMC551627

Purification and characterization of tyrosylprotein sulfotransferase.


Tyrosylprotein sulfotransferase (TPST) is a Golgi membrane enzyme involved in the post-translational modification of secretory and membrane proteins. Here we describe the 140,000-fold purification of this enzyme from bovine adrenal medulla to apparent homogeneity and determine its substrate specificity. The key step in the purification was affinity chromatography on a substrate peptide to which the enzyme bound in the presence of nucleotide cosubstrate. TPST is a 54-50 kd integral membrane glycoprotein. The presence of sialic acid strongly suggests that within the Golgi complex, TPST is localized in the trans-most subcompartment. TPST was found to specifically sulfate tyrosine residues adjacent to acidic amino acids. These results define a major determinant for the specificity of protein sulfation in the trans Golgi.

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  • Almendral JM, Huebsch D, Blundell PA, Macdonald-Bravo H, Bravo R. Cloning and sequence of the human nuclear protein cyclin: homology with DNA-binding proteins. Proc Natl Acad Sci U S A. 1987 Mar;84(6):1575–1579. [PMC free article] [PubMed]
  • Anastasi A, Bertaccini G, Erspamer V. Pharmacological data on phyllokinin (bradykinyl-isoleucyl-tyrosine o-sulphate) and bradykinyl-isoleucyl-tyrosine. Br J Pharmacol Chemother. 1966 Sep;27(3):479–485. [PMC free article] [PubMed]
  • Anderson RG, Pathak RK. Vesicles and cisternae in the trans Golgi apparatus of human fibroblasts are acidic compartments. Cell. 1985 Mar;40(3):635–643. [PubMed]
  • Baeuerle PA, Huttner WB. Tyrosine sulfation is a trans-Golgi-specific protein modification. J Cell Biol. 1987 Dec;105(6 Pt 1):2655–2664. [PMC free article] [PubMed]
  • Barling PM, Palmer DJ, Christie DL. Preparation of desulphated bovine fibrinopeptide B and demonstration of its sulphation in vitro by an enzyme system from neuroblastoma-glioma hybrid cells. Int J Biochem. 1986;18(2):137–141. [PubMed]
  • Bendiak B, Schachter H. Control of glycoprotein synthesis. Purification of UDP-N-acetylglucosamine:alpha-D-mannoside beta 1-2 N-acetylglucosaminyltransferase II from rat liver. J Biol Chem. 1987 Apr 25;262(12):5775–5783. [PubMed]
  • Benedum UM, Lamouroux A, Konecki DS, Rosa P, Hille A, Baeuerle PA, Frank R, Lottspeich F, Mallet J, Huttner WB. The primary structure of human secretogranin I (chromogranin B): comparison with chromogranin A reveals homologous terminal domains and a large intervening variable region. EMBO J. 1987 May;6(5):1203–1211. [PMC free article] [PubMed]
  • Bhavanandan VP, Katlic AW. The interaction of wheat germ agglutinin with sialoglycoproteins. The role of sialic acid. J Biol Chem. 1979 May 25;254(10):4000–4008. [PubMed]
  • Bodanszky M, Martinez J, Priestley GP, Gardner JD, Mutt V. Cholecystokinin (pancreozymin). 4. Synthesis and properties of a biologically active analogue of the C-terminal heptapeptide with epsilon-hydroxynorleucine sulfate replacing tyrosine sulfate. J Med Chem. 1978 Oct;21(10):1030–1035. [PubMed]
  • Brandan E, Hirschberg CB. Purification of rat liver N-heparan-sulfate sulfotransferase. J Biol Chem. 1988 Feb 15;263(5):2417–2422. [PubMed]
  • Cleveland DW, Fischer SG, Kirschner MW, Laemmli UK. Peptide mapping by limited proteolysis in sodium dodecyl sulfate and analysis by gel electrophoresis. J Biol Chem. 1977 Feb 10;252(3):1102–1106. [PubMed]
  • Cooper JA, Gould KL, Cartwright CA, Hunter T. Tyr527 is phosphorylated in pp60c-src: implications for regulation. Science. 1986 Mar 21;231(4744):1431–1434. [PubMed]
  • Daley LA, Renosto F, Segel IH. ATP sulfurylase-dependent assays for inorganic pyrophosphate: applications to determining the equilibrium constant and reverse direction kinetics of the pyrophosphatase reaction, magnesium binding to orthophosphate, and unknown concentrations of pyrophosphate. Anal Biochem. 1986 Sep;157(2):385–395. [PubMed]
  • Elices MJ, Blake DA, Goldstein IJ. Purification and characterization of a UDP-Gal:beta-D-Gal(1,4)-D-GlcNAc alpha(1,3)-galactosyltransferase from Ehrlich ascites tumor cells. J Biol Chem. 1986 May 5;261(13):6064–6072. [PubMed]
  • Eng J, Gubler U, Raufman JP, Chang M, Hulmes JD, Pan YC, Yalow RS. Cholecystokinin-associated COOH-terminal peptides are fully sulfated in pig brain. Proc Natl Acad Sci U S A. 1986 May;83(9):2832–2835. [PMC free article] [PubMed]
  • Fessler LI, Brosh S, Chapin S, Fessler JH. Tyrosine sulfation in precursors of collagen V. J Biol Chem. 1986 Apr 15;261(11):5034–5040. [PubMed]
  • Fessler LI, Chapin S, Brosh S, Fessler JH. Intracellular transport and tyrosine sulfation of procollagens V. Eur J Biochem. 1986 Aug 1;158(3):511–518. [PubMed]
  • Friederich E, Fritz HJ, Huttner WB. Inhibition of tyrosine sulfation in the trans-Golgi retards the transport of a constitutively secreted protein to the cell surface. J Cell Biol. 1988 Nov;107(5):1655–1667. [PMC free article] [PubMed]
  • Fukui S, Numata Y, Yamashina I. Comparison of protein sulfation in control and virus-transformed baby hamster kidney cells. J Biochem. 1984 Dec;96(6):1783–1788. [PubMed]
  • Heukeshoven J, Dernick R. Improved silver staining procedure for fast staining in PhastSystem Development Unit. I. Staining of sodium dodecyl sulfate gels. Electrophoresis. 1988 Jan;9(1):28–32. [PubMed]
  • Hirose S, Oda K, Ikehara Y. Tyrosine O-sulfation of the fibrinogen gamma B chain in primary cultures of rat hepatocytes. J Biol Chem. 1988 May 25;263(15):7426–7430. [PubMed]
  • Hortin GL, Farries TC, Graham JP, Atkinson JP. Sulfation of tyrosine residues increases activity of the fourth component of complement. Proc Natl Acad Sci U S A. 1989 Feb;86(4):1338–1342. [PMC free article] [PubMed]
  • Hortin G, Folz R, Gordon JI, Strauss AW. Characterization of sites of tyrosine sulfation in proteins and criteria for predicting their occurrence. Biochem Biophys Res Commun. 1986 Nov 26;141(1):326–333. [PubMed]
  • Huttner WB. Tyrosine sulfation and the secretory pathway. Annu Rev Physiol. 1988;50:363–376. [PubMed]
  • Jukkola A, Risteli J, Niemelä O, Risteli L. Incorporation of sulphate into type III procollagen by cultured human fibroblasts. Identification of tyrosine O-sulphate. Eur J Biochem. 1986 Jan 2;154(1):219–224. [PubMed]
  • Laemmli UK. Cleavage of structural proteins during the assembly of the head of bacteriophage T4. Nature. 1970 Aug 15;227(5259):680–685. [PubMed]
  • Lee RW, Huttner WB. Tyrosine-O-sulfated proteins of PC12 pheochromocytoma cells and their sulfation by a tyrosylprotein sulfotransferase. J Biol Chem. 1983 Sep 25;258(18):11326–11334. [PubMed]
  • Lee RW, Huttner WB. (Glu62, Ala30, Tyr8)n serves as high-affinity substrate for tyrosylprotein sulfotransferase: a Golgi enzyme. Proc Natl Acad Sci U S A. 1985 Sep;82(18):6143–6147. [PMC free article] [PubMed]
  • Liu N, Baenziger JU. In vivo and in vitro tyrosine sulfation of a membrane glycoprotein. J Biol Chem. 1986 Jan 15;261(2):856–861. [PubMed]
  • Lotan R, Nicolson GL. Purification of cell membrane glycoproteins by lectin affinity chromatography. Biochim Biophys Acta. 1979 Dec 20;559(4):329–376. [PubMed]
  • Nachman RJ, Holman GM, Cook BJ, Haddon WF, Ling N. Leucosulfakinin-II, a blocked sulfated insect neuropeptide with homology to cholecystokinin and gastrin. Biochem Biophys Res Commun. 1986 Oct 15;140(1):357–364. [PubMed]
  • Nishikawa Y, Pegg W, Paulsen H, Schachter H. Control of glycoprotein synthesis. Purification and characterization of rabbit liver UDP-N-acetylglucosamine:alpha-3-D-mannoside beta-1,2-N-acetylglucosaminyltransferase I. J Biol Chem. 1988 Jun 15;263(17):8270–8281. [PubMed]
  • Pauwels S, Dockray GJ, Walker R. Comparison of the metabolism of sulfated and unsulfated heptadecapeptide gastrin in humans. Gastroenterology. 1987 May;92(5 Pt 1):1220–1225. [PubMed]
  • Pittman DD, Kaufman RJ. Structure-function relationships of factor VIII elucidated through recombinant DNA technology. Thromb Haemost. 1989 Apr 25;61(2):161–165. [PubMed]
  • Rens-Domiano S, Roth JA. Characterization of tyrosylprotein sulfotransferase from rat liver and other tissues. J Biol Chem. 1989 Jan 15;264(2):899–905. [PubMed]
  • Roth J, Taatjes DJ, Lucocq JM, Weinstein J, Paulson JC. Demonstration of an extensive trans-tubular network continuous with the Golgi apparatus stack that may function in glycosylation. Cell. 1985 Nov;43(1):287–295. [PubMed]
  • Sadler JE, Rearick JI, Paulson JC, Hill RL. Purification to homogeneity of a beta-galactoside alpha2 leads to 3 sialyltransferase and partial purification of an alpha-N-acetylgalactosaminide alpha2 leads to 6 sialyltransferase from porcine submaxillary glands. J Biol Chem. 1979 Jun 10;254(11):4434–4442. [PubMed]
  • Schubert D, LaCorbiere M, Saitoh T, Cole G. Characterization of an amyloid beta precursor protein that binds heparin and contains tyrosine sulfate. Proc Natl Acad Sci U S A. 1989 Mar;86(6):2066–2069. [PMC free article] [PubMed]
  • Stone SR, Hofsteenge J. Kinetics of the inhibition of thrombin by hirudin. Biochemistry. 1986 Aug 12;25(16):4622–4628. [PubMed]
  • Stoscheck CM. Protein assay sensitive at nanogram levels. Anal Biochem. 1987 Feb 1;160(2):301–305. [PubMed]
  • Suiko M, Liu MC. Change in binding affinities of 3Y1 secreted fibronectin upon desulfation of tyrosine-O-sulfate. Biochem Biophys Res Commun. 1988 Aug 15;154(3):1094–1098. [PubMed]
  • Tarentino AL, Gómez CM, Plummer TH., Jr Deglycosylation of asparagine-linked glycans by peptide:N-glycosidase F. Biochemistry. 1985 Aug 13;24(17):4665–4671. [PubMed]
  • Trifaró JM, Duerr AC. Isolation and characterization of a Golgi-rich fraction from the adrenal medulla. Biochim Biophys Acta. 1976 Jan 14;421(1):153–167. [PubMed]
  • Van Beveren C, van Straaten F, Curran T, Müller R, Verma IM. Analysis of FBJ-MuSV provirus and c-fos (mouse) gene reveals that viral and cellular fos gene products have different carboxy termini. Cell. 1983 Apr;32(4):1241–1255. [PubMed]
  • Vargas F, Frerot O, Tuong MD, Schwartz JC. Characterization of a tyrosine sulfotransferase in rat brain using cholecystokinin derivatives as acceptors. Biochemistry. 1985 Oct 8;24(21):5938–5943. [PubMed]
  • Vera JC. Measurement of microgram quantities of protein by a generally applicable turbidimetric procedure. Anal Biochem. 1988 Oct;174(1):187–196. [PubMed]
  • Weidemann A, König G, Bunke D, Fischer P, Salbaum JM, Masters CL, Beyreuther K. Identification, biogenesis, and localization of precursors of Alzheimer's disease A4 amyloid protein. Cell. 1989 Apr 7;57(1):115–126. [PubMed]
  • Weinstein J, de Souza-e-Silva U, Paulson JC. Purification of a Gal beta 1 to 4GlcNAc alpha 2 to 6 sialyltransferase and a Gal beta 1 to 3(4)GlcNAc alpha 2 to 3 sialyltransferase to homogeneity from rat liver. J Biol Chem. 1982 Nov 25;257(22):13835–13844. [PubMed]

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