Polysialyltransferase-1 autopolysialylation is not requisite for polysialylation of neural cell adhesion molecule.
Polysialyltransferase-1 (PST; ST8Sia IV) is one of the alpha2, 8-polysialyltransferases responsible for the polysialylation of the neural cell adhesion molecule (NCAM). The presence of polysialic acid on NCAM has been shown to modulate cell-cell and cell-matrix interactions. We previously reported that the PST enzyme itself is modified by alpha2,8-linked polysialic acid chains in vivo. To understand the role of autopolysialylation in PST enzymatic activity, we employed a mutagenesis approach. We found that PST is modified by five Asn-linked oligosaccharides and that the vast majority of the polysialic acid is found on the oligosaccharide modifying Asn-74. In addition, the presence of the oligosaccharide on Asn-119 appeared to be required for folding of PST into an active enzyme. Co-expression of the PST Asn mutants with NCAM demonstrated that autopolysialylation is not required for PST polysialyltransferase activity. Notably, catalytically active, non-autopolysialylated PST does not polysialylate any endogenous COS-1 cell proteins, highlighting the protein specificity of polysialylation. Immunoblot analyses of NCAM polysialylation by polysialylated and non-autopolysialylated PST suggests that the NCAM is polysialylated to a higher degree by autopolysialylated PST. We conclude that autopolysialylation of PST is not required for, but does enhance, NCAM polysialylation.
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B E
BE
Department of Biochemistry and Molecular Biology, University of Illinois College of Medicine, Chicago, Illinois 60612, USA.
Tao
K
K
Colley
K J
KJ
D000818:Animals; D001216:Asparagine / Q000737:chemistry; D019556:COS Cells; D005455:Fluorescent Antibody Technique; D015151:Immunoblotting; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D019006:Neural Cell Adhesion Molecules / Q000378:metabolism*; D011233:Precipitin Tests; D012794:Sialic Acids / Q000737:chemistry; D012799:Sialyltransferases / Q000235:genetics / Q000378:metabolism*
D019006:Neural Cell Adhesion Molecules; D012794:Sialic Acids; C021319:polysialic acid; D001216:Asparagine; D012799:Sialyltransferases