HNK-1抗原の生合成に関与するグルクロン酸転移酵素とその機能に関する研究

HNK-1抗原生物合成相关葡萄糖醛酸转移酶及其功能研究

基本信息

批准号：
12470497
负责人：
KAWASAKI Toshisuke
金额：
$ 10.3万
依托单位：
KYOTO UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (B)
财政年份：
2000
资助国家：
日本
起止时间：
2000 至 2001
项目状态：
已结题

项目摘要

In our previous studies, we succeeded in cloning of two kind of glucuronyltransferase (GlcAT-P and GlcAT-S) cDNAs that are responsible for the biosynthesis of the HNK-1 epitope and studied the functions of the HNK-1 epitope using these cDNAs as molecular probes. In this study, we have cloned the human GlcAT-P gene for the first time and revealed that the en-zyme is a type II membrane protein consisting of 334 amino acids and the amino acid se-quence of the catalytic region is 98.2% identical to that of rat GlcAT-P but is different from the latter in the 13 amino acid shorter C-terminal cytoplasmic domain. The human GlcAT-P gene was mapped to 11q25, which is syntenic with the mouse GlcAT-P locus, the A4 region of chromosome 9. The acceptor specificity of a rat brain glucuronyltransferase, GlcAT-P,was investigated using asialoorosomucoid as a model acceptor substrate. The enzyme trans-ferred glucuronic acid to bi-, tri-, and tetra-antennary complex type sugar chains, with almost equal efficiency, indicating that the enzyme has no preference as to the number of acceptor sugar branches. The GlcAT-P is highly specific for the terminal W-acetyllactosamine structure and no glucuronic acid was incorporated into a Galbl-3GlcNAc branch. The GlcAT-P transferred glucuronic acid to the galactose residues in each N-acetyllactosamine residue of the tetra-antennary oligosaccharide chains with different efficiencies and most preferentially to those in the Galbl-4GlcNAcbl-4Manal-3 branch. With regards to the membrane phospholipid requirement of GlcAT-P, we demonstrated that phosphatidyl inositol is required for the GlcAT-P activity to glycolipid substrate, paragloboside.

在我们先前的研究中，我们成功地克隆了两种葡萄糖lcunylranyltransferase（Glcat-P和Glcat-S）CDNA，这些CDNA负责HNK-1表位的生物合成，并研究了将HNK-1表位的功能以这些CDNAS作为分子探针的功能。在这项研究中，我们首次克隆了人Glcat-P基因，并揭示了Zyme是由334个氨基酸组成的II型膜蛋白，催化区域的氨基酸分离是98.2％的98.2％，与大鼠Glcat-P相似，但与后者在13 Amino Cider Cyer cyll cylastrasin中不同。将人Glcat-P基因映射到11q25，该基因与小鼠Glcat-P基因座（染色体9的A4区域）同步。使用AsialOorosomocoid作为模型受体底物研究了大鼠脑葡萄糖酰基转移酶的受体特异性Glcat-P，Glcat-P，Glcat-P，Glcat-P，Glcat-P。酶将葡萄糖酸转移到双，三 - 甲状腺复合型糖链，几乎相等，表明该酶对受体糖分支的数量不偏爱。 GLCAT-P高度特异性对于末端W-乙酰乳糖胺结构，未将葡萄糖酸掺入Galbl-3GLCNAC分支中。 GLCAT-P将葡萄糖酸转移到二甲甲胺残基中的半乳糖残基中，其甲基酸糖链的每个N-环甲酸残基具有不同的效率，并且最优先地比Galbl-4GlCNACBL-4MANAL-4MANAL-4MANAL-MANAL-3分支中最优先。关于GLCAT-P的膜磷脂需求，我们证明了Glcat-P活性对糖脂底物，paragloboboside是必需的。