31 P nuclear magnetic resonance studies on the phosphoglycopeptides obtained from rat brain glycoprotein

Davis, Leonard G.; Costello, Anthony J.; Javaid, Javaid I.; Brunngraber, Eric G.

doi:10.1016/0014-5793(76)80615-1

Cited by 10 publications

(4 citation statements)

References 15 publications

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“…However, there was precedent for this structure in yeast phosphomannans (33). There was also evidence for phosphomannosyl glycoprotein from rat brain, with the phosphate tentatively identified as being present on a secondary alcohol group of the mannose (34,35). The direct evidence that Man6P is present in acid hydrolases and plays a role in directing them to lysosomes raises a number of new and interesting questions.…”

Section: Discussionmentioning

confidence: 99%

Enzymatic identification of mannose 6-phosphate on the recognition marker for receptor-mediated pinocytosis of beta-glucuronidase by human fibroblasts.

Natowicz¹,

Maggie²,

Lowry³

et al. 1979

Proc. Natl. Acad. Sci. U.S.A.

208

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Human fB-glucuronidase (fl-D-glucuronide glucuronosohydrolase, EC 3.2.1.31), like many other glycopr tein lysosomal hydrolases, is subject to receptor-mediated endocytosis by fibroblasts. Prior work demonstrated charge heterogeneity in P-glucuronidase and showed that high-uptake forms are more acidic than slowly internalized forms. Considerable indirect evidence implicated mannose 6-phosphate as an essential part of the recognition marker on high-uptake enzyme forms. Here we report the purification of fl-glucuronidase from human spleen and demonstrate enzymatically that mannose 6-phosphate is released on acid hydrolysis of pure enzyme. Furthermore, the mannose 6-phosphate content of the enzyme varies directly with its susceptibility to pinocytosis by fi roblasts. Enzyme forms resolved by CM-Sephadex chromatography differed over an 18-fold range in uptake rate and in mannose 6-phosphate content. The most acidic forms had 4.4 mol of mannose 6-phosphate per mol of enzyme. The mannose 6-phosphate was released from the enz me b treatment with endoglycosidase H with concomitant loss of susceptibility to adsorptive endocytosis. Thus, these studies provide direct evidence that mannose 6-phosphate is present on high-uptake enzyme forms,.that it is present in the recognition marker for uptake, and that it is present on oligosaccharide that is released by endoglycosidase H.

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Section: Discussionmentioning

confidence: 99%

Enzymatic identification of mannose 6-phosphate on the recognition marker for receptor-mediated pinocytosis of beta-glucuronidase by human fibroblasts.

Natowicz¹,

Maggie²,

Lowry³

et al. 1979

Proc. Natl. Acad. Sci. U.S.A.

208

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“…This may suggest that either phosphorylated mannose residues adjacent to N-acetylglucosamine residues inhibit the action of P-N-acetylglucosaminidase or that these N-acetylglucosamine residues are phosphorylated. A phosphorylated glycoprotein has been prepared from rat brain [26], from which a glycopeptide with an average molecular weight of 2200 was isolated consisting of mannose and N-acetylglucosamine and bearing the phosphate group at a secondary alcoholic group of the carbohydrates [27].…”

Section: Discussionmentioning

confidence: 99%

Isolation and Characterization of Phosphorylated Oligosaccharides from α‐N‐Acetylglucosaminidase that Are Recognized by Cell‐Surface Receptors

Figura

Klein

1979

European Journal of Biochemistry

101

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Adsorptive endocytosis of lysosomal enzymes by fibroblasts and hepatocytes involves binding to cell surface receptors that recognize on lysosomal enzymes a phosphorylated carbohydrate, most likely a mannose 6-phosphate residue Proc. Nut1 Acud. Sci. U.S.A. 74, 2026Sci. U.S.A. 74, -2030Ullrich et al. (1978) Hoppe-Seyler's Z. Physiol. Chem. 359, 1591. Loss of a-Nacetylglucosaminidase endocytosis after treatment with endoglucosaminidase H indicated that the recognition site of a-N-acetylglucosaminidase is located on N-glycosidically linked oligosaccharides of the high mannose type. Acidic oligosaccharides with an average molecular weight of 2200 were liberated from wN-acetylglucosaminidase by endoglucosaminidase H. These oligosaccharides were susceptible to degradation by alkaline phosphatase, a-mannosidase and P-N-acetylglucosaminidase. At the non-reducing terminal these oligosaccharides bear phosphorylated mannose and/or N-acetylgiucosamine residues.Cultured cells may internalize lysosomal enzymes by a receptor-mediated endocytosis (for review see [l 3). Cell surface receptors on fibroblasts recognize on lysosomal enzymes a phosphorylated carbohydrate, most likely a mannose 6-phosphate residue [2 -51. Hepatocytes possess receptors with the same ligand specificity [6,7]. In addition, hepatocytes interact with terminal galactose [7 -The present results describe the isolation and characterization of phosphorylated oligosaccharides from human urine a-N-acetylglucosaminidase. and mannose [7] residues on lysosomal enzymes; whereas macrophages possess a receptor system that interacts both with N-acetylglucosamine and with mannose residues on lysosomal enzymes [lo -131. MATERIALS AND METHODS MaterialsEvidence for the presence of a phosphorylated carbohydrate on lysosomal enzymes has been so far only indirect : (a) mannose 6-phosphate, phosphorylated mannans and bovine serum albumin substituted with mannose 6-phosphate are competitive inhibitors for endocytosis of lysosomal enzymes by fibroblasts [2 -5,14,15] and (b) periodate oxidation and alkaline phosphatase treatment destroy the recognition site on lysosomal enzymes [2 -5,161. NaB[3H4] (specific activity 7 Ci/mmol) was obtained from Amersham-Buchler (Braunschweig). Sephadex G-25 fine and concanavalin-A -Sepharose were from Pharmacia (Uppsala). Endoglucosaminidase H and D were from Miles (Frankfurt), P-N-acetylglucosaminidase (jack bean, specific activity 38 U/mg protein) from Sigma Chem. (Miinchen), alkaline phosphatase (Escherichia coli, specific activity 40 Ujmg) from Boehringer Mannheim (Mannheim), neuraminidase (Vibrio cholerae, specific activity 4 Ujmg) from Behringwerke (Marburg). P-Galactosidase and P-Nacetylglucosaminidase from Diplococcus pneumoniue were those m-eviouslv described 117,201. Purified substrate per min at 37 "C.

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“…N.m.r. studies [20] revealed the absence of phosphoserine, phospholipid, and diester phosphate linkages. None of the label could be extracted into an organic phase.…”

Section: Resultsmentioning

confidence: 98%

“…The phosphorylated, Con A-binding, mannose rich glycopeptides appear to be the only phosphate-containing heteropolysaccharide chains associated with brain glycoproteins [20]. Structural, metabolic, subcellular and anatomic distribution studies pertaining to these phosphoglycoproteins is under investigation.…”

Section: Discussionmentioning

confidence: 99%

Identification of phosphoglycoproteins obtained from rat brain

1976

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31 P nuclear magnetic resonance studies on the phosphoglycopeptides obtained from rat brain glycoprotein

Cited by 10 publications

References 15 publications

Enzymatic identification of mannose 6-phosphate on the recognition marker for receptor-mediated pinocytosis of beta-glucuronidase by human fibroblasts.

Enzymatic identification of mannose 6-phosphate on the recognition marker for receptor-mediated pinocytosis of beta-glucuronidase by human fibroblasts.

Isolation and Characterization of Phosphorylated Oligosaccharides from α‐N‐Acetylglucosaminidase that Are Recognized by Cell‐Surface Receptors

Identification of phosphoglycoproteins obtained from rat brain

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