Glycoblology vol 2 no. 2 pp. 119-125, 1992
Purification and characterization of human lymphoblast N-acetylglucosamine-1-phosphotransferase
Ke-Wei Zhao, Richard Yeh and Arnold L.Miller1 Department of Neurosciences, 0624, University of California, San Diego School of Medicine, La Jolla, CA 92093, USA 'To whom correspondence should be addressed
iV-Acetylglucosamine-1-phosphotransferase (GlcNAcPTase) was solubilized with 2% Tergitol NP-10 from cultured human lymphoblast cells and purified 3840-fold with 14% recovery using lentil lectin-Sepharose 4B, DEAE-Sephacel and Sephacryl S-400 chromatographies. The partially purified enzyme requires the non-ionic detergent Tergitol NP-10 and a divalent cation, Mn2"^ or Mg 2+ , for its activity and exhibits an optimal pH at 7.2-7.5 in Tris-maleate buffer. Kinetic studies demonstrated an apparent Km of 24 /iM for the donor UDP-N-acetylglucosamine and of 117 mM for the artificial acceptor a-methylmannoside. The GlcNAcPTase is inhibited by UDP and UDP-glucose, and by negatively charged phospholipids including phosphatidylserine, phosphatidylglycerol and phosphatidic acid. The apparent mol. wt of the human lymphoblast GlcNAcPTase is ~ 1000 kDa, which is analogous to that reported for the partially purified enzyme from rat liver (Waheed et at., 1982). Key words: /V-acetylglucosamine-1-phosphotransferase/lysosomal enzyme targeting/mannose 6-phosphate/oligosaccharide phosphorylation
be dependent on a specific amino acid sequence, but on a tertiary conformation involving a lysine residue and a discontinuous peptide loop structure associated with lysosomal enzymes (Lang et ai, 1984; Baranski et al., 1990). The importance of GlcNAcPTase in regulating intracellular trafficking of lysosomal enzymes is seen in the altered targeting that occurs for many of these enzymes in cells of mesenchymal origin in I-cell disease and pseudo-Hurler polydystrophy (Nolan and Sly, 1989). These two related autosomal recessive disorders occur due to a severe decrease or absence of GlcNAcPTase activity (Hasilik et at, 1981; Reitman etai, 1981). Although GlcNAcPTase plays a key role in the biosynthesis and targeting of lysosomal enzymes to lysosomes, little information is currently available concerning its structure and the gene product(s) that control its expression. An understanding of the structure and interaction of the gene product(s) becomes of paramount importance when one evaluates the reasons for the genetic complementation groups reported to occur within and between these two disorders (Gravel et al., 1981; Honey etal, 1981, 1982; Shows etai, 1982; Mueller et al., 1983; Little et al., 1986). A purified or partially purified GlcNAcPTase is essential for these aforementioned studies. Published results concerning the purification and characterization of the GlcNAcPTase have been minimal (Waheed et al., 1982; Reitman et al., 1984), indicating the difficult nature of purifying this enzyme. The present study represents the first report describing the procedure for obtaining partially purified human lymphoblast GlcNAcPTase and its characterization. Preliminary reports of these findings have appeared elsewhere (Zhao etal., 1990a, b).
Introduction The ability of many lysosomal enzymes to segregate to lysosomes is dependent on possessing a targeting signal, mannose 6-phosphate (Kornfeld, 1987). The generation of this signal occurs through the action of a key enzyme, UDP/V-acetylglucosamine (UDP-GlcNAc): lysosomal enzyme N-acetylglucosamine 1-phosphotransferase (GlcNAcPTase, E.C. 2.7.8.17) activity (Reitman and Kornfeld, 1981a). This enzyme transfers a-N-acetylglucosamine-l -phosphate from UDP-GlcNAc to selected mannose residues on high-mannose or hybrid oligosaccharide chains on lysosomal enzymes (Tabas and Kornfeld, 1980; Varki and Kornfeld, 1983). The terminal N-acetylglucosamine residues of the resulting structure are subsequently removed by an a-iV-acetylglucosamine-1-phosphodiester N-acetylglucosaminidase (phosphodiester glycosidase, E.C. 3.1.4.45) exposing one or more phosphomonoester(s), mannose 6-phosphate, which is required for recognition of the lysosomal enzymes by a specific transport receptor for subsequent targeting to lysosomes (Varki and Kornfeld, 1980, 1981; Sly and Fischer, 1982). The efficiency of phosphorylation for lysosomal enzymes is at least 100-fold better compared to glycopeptides containing identical oligosaccharide chains (Reitman and Kornfeld, 1981b). The basis for the selectivity of the GlcNAcPTase in phosphorylating only the mannose residues present on lysosomal enzymes appears not to © Oxford University Press
Results Purification of GlcNAcPTase The human lymphoblast GlcNAcPTase was solubilized in the non-ionic detergent Tergitol NP-10, and subsequently purified ~ 3800-fold using a sequence of lentil lectin-Sepharose 4B affinity chromatography, DEAE-Sephacel anion exchange chromatography and Sephacryl S-400 gel filtration. The results of a typical purification are summarized in Table I. Solubilization of GlcNAcPTase. The initial homogenization of lymphoblasts in buffered isotonic sucrose (buffer 1) removed 10-40% of the total protein, whereas the loss of GlcNAcPTase activity was minimal (mannoside /31-2 A'-acetylglucosaminyltransferase II from rat liver. J. Biol. Chem., 262, 5784-5790. Beyer.T.A , SadlerJ.E., Rearick.J.I., PaulsonJ.C. and Hill.R.L. (1981) Glycosyltransferases and their use in assessing oligosacchande structure and structure-function relationships. Adv. Enzymol., 52, 23-175. Conover.J.H., Hathaway.P., Glade.P.R. and Hirschhorn,K. (1970) Persistence of phosphoglucomutase (PGM) polymorphism in long-term lymphoid lines. Proc. Soc. Exp. Biol. Med., 133, 750-753 Gravel,R.A., Gravel,Y.L., Miller.A.L. and LowdenJ.A. (1981) Genetic complementation analysis of I-cell disease and pseudo-Hurler polydystrophy. In Callahan.J.W. and Lowden.J. A. (eds), Lysosomes and Lysosomal Storage Diseases. Raven Press, New York, pp. 289-298. Harms,E., Kartenbach.J., Darai.G. and Schneider,.) (1981) Purification and characterization of human lysosomes from EB-virus transformed lymphoblasts. Exp. Cell Res., 131, 251-266. Hasilik.A., Waheed.A. and von Figura.K. (1981) Enzymatic phosphorylarion of lysosomal enzymes in the presence of UDP-A'-acetylglucosamine: absence of the activity in I-cell fibroblasts. Biochem. Biophys. Res. Common., 98, 712-767. Honey,N K , Miller.A.L. and Shows.T.B (1981) The mucolipidoses: Identification by abnormal electrpphoretic patterns of lysosomal hydrolases Am. J. Med. Genet., 9, 239-253. Honey,N.K., Mueller.O.T., Honey.N.K., Wnght.C.E. and Miller.A.L (1982) Mucolipidosis in is genetically heterogeneous. Proc. Natl. Acad. Sci. USA, 79, 7240-7244. Komfeld.S. (1987) Trafficking of lysosomal enzymes. FASEBJ., 1, 462^68. Lang.L., Reitman.M., TangJ., Roberts,M.R. and Kornfeld.S (1984) Lysosomal enzyme phosphorylation: recognition of a protein-dependent determinant allows specific phosphorylation of oligosaccharides present on lysosomal enzymes. J. Biol Chem., 259, 14663-14671. Little.L., Alcouloumre.M., Drotar.A.M., Herman,S., Robertson,R., Yen, R.Y. and Miller,A L. (1987) Properties of A'-acetylglucosamine 1-phosphotransferase from human lymphoblasts. Biochem. J., 248, 151-159 Little.L.E., Mueller,O.T., Honey,N.K., Shows,T.B. and Miller.A.L. (1986) Heterogeneity of A'-acetylglucosamine 1-phosphotransferase within mucolipidosis m. J. Biol. Chem., 261, 733-738. Lowry.O.H., Rosebrough.M.J., Farr,A.L. and Randall.R.J. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem., 193, 265—275. Mueller.OT., Honey.N.K., Little.L.E., Miller.A.L. and Shows,T.B. (1983) Mucolipidosis n and in. The genetic relationships between two disorders of lysosomal enzyme biosynthesis. J Gin. Invest., 72, 1016—1023 Newby,A.C. and Chrambach.A. (1979) Disaggregation of adenylate cyclase during polyacrylamide-gel electrophoresis in mixtures of ionk and non-ionic detergents. Biochem. J., 177, 623-630. Nishikawa.Y., Pegg.W., Paulsen.H. and Schachter,H. (1988) Control of glycoprotein synthesis: purification and characterization of rabbit liver UDP-A7acetylglucosamine:a-3-r>mannoside B-1,2-A'-acetylglucosaminyltransferase I. J. Biol. Chem., 263, 8270-8281 Nolan.C.M. and Sly.W.S. (1989) I-cell disease and pseudo-Hurier polydystrophy. In Sriver.C.R., Beaudet.A.L., Sly.W.S. and Valle.D. (eds), The Metabolic Basis of Inherited Disease 6th edn. McGraw-Hill Inc., New York, pp. 1589-1601. Reitman.M.L. and Kornfeld.S. (1981a) UDP-A'-acetylglucosamine: glycoprotein A'-acetylglucosamine 1-phosphotransferase. Proposed enzyme for the phosphorylation of the high mannose oligosacchande units of lysosomal enzymes. J. Biol. Chem., 256, 4275-4281. Reitman.M.L. and Komfeld.S. (1981b) Lysosomal enzyme targeting: Af-acetylglucosaminyl-phosphotransferase selectively phosphorylates native lysosomal enzymes. /. Biol. Chem., 256, 11977-11980. Reitman.M.L., Varki.A. and Kornfeld.S. (1981) Fibroblasts from patients with I-cell disease and pseudo-Hurler polydystrophy are deficient in UDP-A'acetylglucosamine: glycoprotein A'-acetylglucosaminyl-phosphotransferasc activity. J. Clin. Invest., 67, 1574-1579. Reitman.M.L., Lang.L. and Kornfeld.S. (1984) UDP-A'-acetylglucosamine: lysosomal enzyme A'-acetylglucosamine 1-phosphotransferase. Methods Enzymol., 107, 163-172.
Purification and properties of GkNAc pbosphotransferase Shows.T.B., Mueller,O.T., Honey,N.K., Wright,C.E. and Miller.A.L. (1982) Genetic heterogeneity of I-cell disease is demonstrated by complementation of lysosomal enzyme processing mutants. Am. J. Med. Genet., 12, 343-353. Sly.W.S. and Fischer.D.D. (1982) The phosphomannosyl recognition system for intracellular and intercellular transport of lysosomal enzymes. /. Cell Biochem., 18, 67-85. Tabas, I. and Kornfeld, S. (1980) Biosynthetic intermediates of |3-glucuronidase contain high mannose oligosacchandes with blocked phosphate residues. J. Biol. Chem., 255, 6633-6639. Varki.A. and Komfeld.S. (1980) Identification of a rat liver a-N-acetylglucosaminyl phosphodiesterase capable of removing 'blocking' a-N-acetylglucosamine residues from phosphorylated high mannose oligosacchandes of lysosomal enzymes. J. Bid. Chem., 255, 8398-8401. Varki.A. and Komfeld.S. (1981) Purification and characterization of rat liver a-W-acetylglucosaminyl phosphodiesterase. J. Biol. Chem., 256, 9937-9943. Varki.A. and Kornfeld.S. (1983) The spectrum of anionic oligosaccharides released by endo-/3-W-acetylglucosaminidase H from glycoproteins J. Biol. Chem., 258, 2808-2818. Waheed.A., Hailik.A. and von Figure,K. (1982) UDP-iV-acetylglucosamme. lysosomal enzyme precursor A'-acetylglucosamine 1-phosphotransferase. Partial purification and characterization of the rat liver Golgi enzyme. J. Biol. Chem., 257, 12322-12331. Zhao.K., Yeh.R. and Miller.A.L. (1990a) Human lymphoblast N-acetylglucosamine-1-phosphotransferase. partial purification ami characterization. FASEB J., 4, A1980. Zhao,K., Yeh.R. and Miller.A.L. (1990b) Properties of highly purified human lymphoblast iV-acetylglucosamine- 1-phosphotransferase. Gtycoconjugate J., 7, 497. Zhao.K., Yeh.R and Miller.A.L (1991) Punfication of JV-acetylglucosamine-1-phosphotransferase from human lymphoblasts using affinity chromatography. Gtycoconjugate J., 8, 267-268. Received on September 20, 1991; accepted on October 14, 1991
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Purification and characterization of human lymphoblast N-acetylglucosamine-1-phosphotransferase
Ke-Wei Zhao, Richard Yeh and Arnold L.Miller1 Department of Neurosciences, 0624, University of California, San Diego School of Medicine, La Jolla, CA 92093, USA 'To whom correspondence should be addressed
iV-Acetylglucosamine-1-phosphotransferase (GlcNAcPTase) was solubilized with 2% Tergitol NP-10 from cultured human lymphoblast cells and purified 3840-fold with 14% recovery using lentil lectin-Sepharose 4B, DEAE-Sephacel and Sephacryl S-400 chromatographies. The partially purified enzyme requires the non-ionic detergent Tergitol NP-10 and a divalent cation, Mn2"^ or Mg 2+ , for its activity and exhibits an optimal pH at 7.2-7.5 in Tris-maleate buffer. Kinetic studies demonstrated an apparent Km of 24 /iM for the donor UDP-N-acetylglucosamine and of 117 mM for the artificial acceptor a-methylmannoside. The GlcNAcPTase is inhibited by UDP and UDP-glucose, and by negatively charged phospholipids including phosphatidylserine, phosphatidylglycerol and phosphatidic acid. The apparent mol. wt of the human lymphoblast GlcNAcPTase is ~ 1000 kDa, which is analogous to that reported for the partially purified enzyme from rat liver (Waheed et at., 1982). Key words: /V-acetylglucosamine-1-phosphotransferase/lysosomal enzyme targeting/mannose 6-phosphate/oligosaccharide phosphorylation
be dependent on a specific amino acid sequence, but on a tertiary conformation involving a lysine residue and a discontinuous peptide loop structure associated with lysosomal enzymes (Lang et ai, 1984; Baranski et al., 1990). The importance of GlcNAcPTase in regulating intracellular trafficking of lysosomal enzymes is seen in the altered targeting that occurs for many of these enzymes in cells of mesenchymal origin in I-cell disease and pseudo-Hurler polydystrophy (Nolan and Sly, 1989). These two related autosomal recessive disorders occur due to a severe decrease or absence of GlcNAcPTase activity (Hasilik et at, 1981; Reitman etai, 1981). Although GlcNAcPTase plays a key role in the biosynthesis and targeting of lysosomal enzymes to lysosomes, little information is currently available concerning its structure and the gene product(s) that control its expression. An understanding of the structure and interaction of the gene product(s) becomes of paramount importance when one evaluates the reasons for the genetic complementation groups reported to occur within and between these two disorders (Gravel et al., 1981; Honey etal, 1981, 1982; Shows etai, 1982; Mueller et al., 1983; Little et al., 1986). A purified or partially purified GlcNAcPTase is essential for these aforementioned studies. Published results concerning the purification and characterization of the GlcNAcPTase have been minimal (Waheed et al., 1982; Reitman et al., 1984), indicating the difficult nature of purifying this enzyme. The present study represents the first report describing the procedure for obtaining partially purified human lymphoblast GlcNAcPTase and its characterization. Preliminary reports of these findings have appeared elsewhere (Zhao etal., 1990a, b).
Introduction The ability of many lysosomal enzymes to segregate to lysosomes is dependent on possessing a targeting signal, mannose 6-phosphate (Kornfeld, 1987). The generation of this signal occurs through the action of a key enzyme, UDP/V-acetylglucosamine (UDP-GlcNAc): lysosomal enzyme N-acetylglucosamine 1-phosphotransferase (GlcNAcPTase, E.C. 2.7.8.17) activity (Reitman and Kornfeld, 1981a). This enzyme transfers a-N-acetylglucosamine-l -phosphate from UDP-GlcNAc to selected mannose residues on high-mannose or hybrid oligosaccharide chains on lysosomal enzymes (Tabas and Kornfeld, 1980; Varki and Kornfeld, 1983). The terminal N-acetylglucosamine residues of the resulting structure are subsequently removed by an a-iV-acetylglucosamine-1-phosphodiester N-acetylglucosaminidase (phosphodiester glycosidase, E.C. 3.1.4.45) exposing one or more phosphomonoester(s), mannose 6-phosphate, which is required for recognition of the lysosomal enzymes by a specific transport receptor for subsequent targeting to lysosomes (Varki and Kornfeld, 1980, 1981; Sly and Fischer, 1982). The efficiency of phosphorylation for lysosomal enzymes is at least 100-fold better compared to glycopeptides containing identical oligosaccharide chains (Reitman and Kornfeld, 1981b). The basis for the selectivity of the GlcNAcPTase in phosphorylating only the mannose residues present on lysosomal enzymes appears not to © Oxford University Press
Results Purification of GlcNAcPTase The human lymphoblast GlcNAcPTase was solubilized in the non-ionic detergent Tergitol NP-10, and subsequently purified ~ 3800-fold using a sequence of lentil lectin-Sepharose 4B affinity chromatography, DEAE-Sephacel anion exchange chromatography and Sephacryl S-400 gel filtration. The results of a typical purification are summarized in Table I. Solubilization of GlcNAcPTase. The initial homogenization of lymphoblasts in buffered isotonic sucrose (buffer 1) removed 10-40% of the total protein, whereas the loss of GlcNAcPTase activity was minimal (mannoside /31-2 A'-acetylglucosaminyltransferase II from rat liver. J. Biol. Chem., 262, 5784-5790. Beyer.T.A , SadlerJ.E., Rearick.J.I., PaulsonJ.C. and Hill.R.L. (1981) Glycosyltransferases and their use in assessing oligosacchande structure and structure-function relationships. Adv. Enzymol., 52, 23-175. Conover.J.H., Hathaway.P., Glade.P.R. and Hirschhorn,K. (1970) Persistence of phosphoglucomutase (PGM) polymorphism in long-term lymphoid lines. Proc. Soc. Exp. Biol. Med., 133, 750-753 Gravel,R.A., Gravel,Y.L., Miller.A.L. and LowdenJ.A. (1981) Genetic complementation analysis of I-cell disease and pseudo-Hurler polydystrophy. In Callahan.J.W. and Lowden.J. A. (eds), Lysosomes and Lysosomal Storage Diseases. Raven Press, New York, pp. 289-298. Harms,E., Kartenbach.J., Darai.G. and Schneider,.) (1981) Purification and characterization of human lysosomes from EB-virus transformed lymphoblasts. Exp. Cell Res., 131, 251-266. Hasilik.A., Waheed.A. and von Figura.K. (1981) Enzymatic phosphorylarion of lysosomal enzymes in the presence of UDP-A'-acetylglucosamine: absence of the activity in I-cell fibroblasts. Biochem. Biophys. Res. Common., 98, 712-767. Honey,N K , Miller.A.L. and Shows.T.B (1981) The mucolipidoses: Identification by abnormal electrpphoretic patterns of lysosomal hydrolases Am. J. Med. Genet., 9, 239-253. Honey,N.K., Mueller.O.T., Honey.N.K., Wnght.C.E. and Miller.A.L (1982) Mucolipidosis in is genetically heterogeneous. Proc. Natl. Acad. Sci. USA, 79, 7240-7244. Komfeld.S. (1987) Trafficking of lysosomal enzymes. FASEBJ., 1, 462^68. Lang.L., Reitman.M., TangJ., Roberts,M.R. and Kornfeld.S (1984) Lysosomal enzyme phosphorylation: recognition of a protein-dependent determinant allows specific phosphorylation of oligosaccharides present on lysosomal enzymes. J. Biol Chem., 259, 14663-14671. Little.L., Alcouloumre.M., Drotar.A.M., Herman,S., Robertson,R., Yen, R.Y. and Miller,A L. (1987) Properties of A'-acetylglucosamine 1-phosphotransferase from human lymphoblasts. Biochem. J., 248, 151-159 Little.L.E., Mueller,O.T., Honey,N.K., Shows,T.B. and Miller.A.L. (1986) Heterogeneity of A'-acetylglucosamine 1-phosphotransferase within mucolipidosis m. J. Biol. Chem., 261, 733-738. Lowry.O.H., Rosebrough.M.J., Farr,A.L. and Randall.R.J. (1951) Protein measurement with the Folin phenol reagent. J. Biol. Chem., 193, 265—275. Mueller.OT., Honey.N.K., Little.L.E., Miller.A.L. and Shows,T.B. (1983) Mucolipidosis n and in. The genetic relationships between two disorders of lysosomal enzyme biosynthesis. J Gin. Invest., 72, 1016—1023 Newby,A.C. and Chrambach.A. (1979) Disaggregation of adenylate cyclase during polyacrylamide-gel electrophoresis in mixtures of ionk and non-ionic detergents. Biochem. J., 177, 623-630. Nishikawa.Y., Pegg.W., Paulsen.H. and Schachter,H. (1988) Control of glycoprotein synthesis: purification and characterization of rabbit liver UDP-A7acetylglucosamine:a-3-r>mannoside B-1,2-A'-acetylglucosaminyltransferase I. J. Biol. Chem., 263, 8270-8281 Nolan.C.M. and Sly.W.S. (1989) I-cell disease and pseudo-Hurier polydystrophy. In Sriver.C.R., Beaudet.A.L., Sly.W.S. and Valle.D. (eds), The Metabolic Basis of Inherited Disease 6th edn. McGraw-Hill Inc., New York, pp. 1589-1601. Reitman.M.L. and Kornfeld.S. (1981a) UDP-A'-acetylglucosamine: glycoprotein A'-acetylglucosamine 1-phosphotransferase. Proposed enzyme for the phosphorylation of the high mannose oligosacchande units of lysosomal enzymes. J. Biol. Chem., 256, 4275-4281. Reitman.M.L. and Komfeld.S. (1981b) Lysosomal enzyme targeting: Af-acetylglucosaminyl-phosphotransferase selectively phosphorylates native lysosomal enzymes. /. Biol. Chem., 256, 11977-11980. Reitman.M.L., Varki.A. and Kornfeld.S. (1981) Fibroblasts from patients with I-cell disease and pseudo-Hurler polydystrophy are deficient in UDP-A'acetylglucosamine: glycoprotein A'-acetylglucosaminyl-phosphotransferasc activity. J. Clin. Invest., 67, 1574-1579. Reitman.M.L., Lang.L. and Kornfeld.S. (1984) UDP-A'-acetylglucosamine: lysosomal enzyme A'-acetylglucosamine 1-phosphotransferase. Methods Enzymol., 107, 163-172.
Purification and properties of GkNAc pbosphotransferase Shows.T.B., Mueller,O.T., Honey,N.K., Wright,C.E. and Miller.A.L. (1982) Genetic heterogeneity of I-cell disease is demonstrated by complementation of lysosomal enzyme processing mutants. Am. J. Med. Genet., 12, 343-353. Sly.W.S. and Fischer.D.D. (1982) The phosphomannosyl recognition system for intracellular and intercellular transport of lysosomal enzymes. /. Cell Biochem., 18, 67-85. Tabas, I. and Kornfeld, S. (1980) Biosynthetic intermediates of |3-glucuronidase contain high mannose oligosacchandes with blocked phosphate residues. J. Biol. Chem., 255, 6633-6639. Varki.A. and Komfeld.S. (1980) Identification of a rat liver a-N-acetylglucosaminyl phosphodiesterase capable of removing 'blocking' a-N-acetylglucosamine residues from phosphorylated high mannose oligosacchandes of lysosomal enzymes. J. Bid. Chem., 255, 8398-8401. Varki.A. and Komfeld.S. (1981) Purification and characterization of rat liver a-W-acetylglucosaminyl phosphodiesterase. J. Biol. Chem., 256, 9937-9943. Varki.A. and Kornfeld.S. (1983) The spectrum of anionic oligosaccharides released by endo-/3-W-acetylglucosaminidase H from glycoproteins J. Biol. Chem., 258, 2808-2818. Waheed.A., Hailik.A. and von Figure,K. (1982) UDP-iV-acetylglucosamme. lysosomal enzyme precursor A'-acetylglucosamine 1-phosphotransferase. Partial purification and characterization of the rat liver Golgi enzyme. J. Biol. Chem., 257, 12322-12331. Zhao.K., Yeh.R. and Miller.A.L. (1990a) Human lymphoblast N-acetylglucosamine-1-phosphotransferase. partial purification ami characterization. FASEB J., 4, A1980. Zhao,K., Yeh.R. and Miller.A.L. (1990b) Properties of highly purified human lymphoblast iV-acetylglucosamine- 1-phosphotransferase. Gtycoconjugate J., 7, 497. Zhao.K., Yeh.R and Miller.A.L (1991) Punfication of JV-acetylglucosamine-1-phosphotransferase from human lymphoblasts using affinity chromatography. Gtycoconjugate J., 8, 267-268. Received on September 20, 1991; accepted on October 14, 1991
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