Demonstration of frequent occurrence of oligosialic acid units in various glycoproteins

The objective of this research project were to demonstrate frequent ocurrence of oligosialic acid structure in glycoproteins and to elucidate their biological functions.The results follow：1.Development of specific probes for detecting various oligosialyl glycan structures：Three monoclonal antibodies were newly prepared(3G9 recognizing8-O-sulfated Neu5Ac-containing oligosialic acid structures，2A11recognizing Neu5Acα→8Neu5Acα→6Glc，AC-1(originally established by Dr.Keiko Nohara)recognizing oligo-N-glycolneuraminic acid.Based on inhibition experiments and immunohistochemistry using these antibodies，a2A11epitope on sea urchin sperm was shown to mediate sperm-egg interaction.2。Identification and structural and biosynthetic studies of oligosialic acid-containing glycoproteins：Calf fetal serum fetuin，α-I D22-macroglobulin and calf and murine Adipo Q(An Adipocyte Specific Protein)were first identified as si/oligosialic acid glycoproteins.Expression of the oligosialyl structures Adipo Q in an adipocyte differentiating cell line increased as the cells were differentiated.Thus，oligosialic acid expression depends on differentiation.3。Examination of differentiation-dependent change of expression of oligosialic acid structures：Expression of oligosialic acid-containing glycoproteins in human premyelocytoma cell line HL 60 was shown to change before and after differentiation.The expression of the oligosialyl epitope was also shown to change in adipose cell line before and after differentiation。Northern blot analysis using knownα2，8-sialyltransferases cDNAs showed that no of these cDNAs were expressed in the cells，suggesting that novel member of theα2，8-sialyltransferases is expressed in the cell line。

The objective of this research project were to demonstrate frequent ocurrence of oligosialic acid structure in glycoproteins and to elucidate their biological functions. The results follow :1. Development of specific probes for detecting various oligosialyl glycan structures : Three monoclonal antibodies were newly prepared (3G9 recognizing 8-O-sulfated Neu5Ac-containing oligosialic acid structures, 2A11 recognizing Neu5Acα→8Neu5Acα→6Glc, AC-1 (originally established by Dr. Keiko Nohara) recognizing oligo-N-glycolylneuraminic acid. Based on inhibition experiments and immunohistochemistry using these antibodies, a 2A11 epitope on sea urchin sperm was shown to mediate sperm-egg interaction.2. Identification and structural and biosynthetic studies of oligosialic acid-containing glycoproteins : Calf fetal serum fetuin, αィイD22ィエD2-macroglobulin and calf and murine Adipo Q (an adipocyte specific protein) were first identified as si/oligosialic acid glycoproteins. Expression of the oligosialyl structures Adipo Q in an adipocyte differentiating cell line increased as the cells were differentiated. Thus, oligosialic acid expression depends on differentiation.3. Examination of differentiation-dependent change of expression of oligosialic acid structures : Expression of oligosialic acid-containing glycoproteins in human premyelocytoma cell line HL60 was shown to change before and after differentiation. The expression of the oligosialyl epitope was also shown to change in adipose cell line before and after differentiation. Northern blot analysis using known α2,8-sialyltransferases cDNAs showed that no of these cDNAs were expressed in the cells, suggesting that novel member of the α2,8-sialyltransferases is expressed in the cell line.