Ligatin binds phosphohexose residues on acidic hydrolases

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Abstract
Ligatin, a receptor that recognizes phosphorylated sugars, was isolated from plasma membranes of mouse macrophages, rat ileum, and rat brain. Several acidic hydrolases including N-acetyl β-D-glucosaminidase (β-NAG) were solubilized with this receptor. The solubilized β-NAG bound to ligatin in vitro as demonstrated by affinity chromatography using the immobilized receptor. β-N-Acetyl D-glucosaminidase-ligatin complexes were dissociated by low concentrations of mannose 6-phosphate (Man6P) and/or glucose 1-phosphate (Glc 1P). The effectiveness of these two phosphomonosaccharides varied depending on the source of the enzyme: ileal β-NAG-ligatin complexes showed a four-fold preferential dissociation with Man6P; macrophage complexes showed a 160-fold preferential dissociation with Glc 1P. Brain complexes dissociated with nearly equal preference for Man6P and Glc 1P. Heterologous complexes displayed the specificity characteristic of the source of the enzyme regardless of the source of the ligatin. Treatment of the solubilized hydrolases with endoglucosaminidase H released phosphorous-32 label from these enzymes and prevented binding of β-NAG to ligatin. However, treatment of the solubilized hydrolases with alkaline phosphatase reduced the binding of β-NAG to ligatin by no more than 30%. This apparent resistance of β-NAG to dephosphorylation was consistent with the chromatographic behavior on QAE of 3H-labeled acidic oligosaccharides isolated from the solubilized hydrolases. The oligosaccharides that contain phosphorylated hexose were less acidic than phosphomonoesters and were insensitive to alkaline phosphatase until subjected to acid hydrolysis. These results suggested the presence of a phosphodiester on β-NAG analogous to the NAC glucosamine 1 P6 mannose present on β-glucuronidase isolated from mouse lymphoma cells (Tabas I, Kornfeld, S: J Biol Chem 255: 6633, 1980).