GLYCOSYLTRANSFERASES, MUCINS & MACROPHAGE RECEPTORS

糖基转移酶、粘蛋白

• 批准号: 3484576
• 负责人: ROBERT L HILL
• 金额: $ 47.56万
• 依托单位: DUKE UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-09-01 至 1993-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3484576
• 关键词: Kupffer's cell; affinity chromatography; antibody; carbohydrate biosynthesis; carbohydrate structure; chemical binding; complementary DNA; enzyme structure; genetic library; glucose receptor; glycosyltransferase; growth /development; high performance liquid chromatography; immunoelectron microscopy; laboratory mouse; laboratory rabbit; laboratory rat; macrophage; molecular cloning; mucins; oligosaccharides; point mutation; protein engineering; protein sequence; protein structure function; submandibular gland; swine; tissue /cell culture; transposon /insertion element

The structure-function relationships of three groups of proteins that are involved in aspects of glycoconjugate biosynthesis or function will be examined. GLYCOSYLTRANSFERASES that act in the biosynthesis of oligosaccharides in glycoconjugates will be studied in several ways. 1) cDNA expression libraries in lambda gtll will be screened with antibodies to certain transferases and clones containing transferase cDNA will be isolated and sequenced. 2) The cDNA for certain transferases will be inserted into appropriate expression vectors to obtain sufficient transferase for structure/function studies or to obtain specifically modified enzyme by site specific mutagenesis. 3) Two transferases that act in mucin oligosaccharide biosynthesis and have resisted purification will be purified and characterized. The two transferases form the core sequence GAl beta 1,3Ga1NAc-O- Ser/Thr. MUCINS that serve to lubricate the gastrointestinal, respiratory and urogenital tracts and to protect epithelial cells from injury and dehydration will be isolated and further characterized, as follows. 1) The cDNA sequence of porcine submaxillary gland cDNA will be completed and the sequence of genomic DNA for submaxillary mucin examined. In addition, the number of genes for mucin in the porcine genome will be determined. 2) Antibodies to mucin and synthetic polynucleotides based on submaxillary mucin cDNA sequence will be used to determine whether other porcine tissues contain submaxillary mucin-like protein or mRNA. It will also be determined with use of these probes whether human submaxillary apomucin is structurally related to porcine submaxillary apomucin. 3) The exact location of oligosaccharides in mucin will be determined. 4) Mucins from other porcine tissues, such as stomach, small intestine and colon will be isolated and structurally characterized. MACROPHAGE RECEPTORS for oligosaccharides will be examined, especially the receptor found only on rat and mouse Kupffer cells. 1) The sequence of the genomic DNA for rat Kupffer cells will be determined. 2) The expression of the receptor during macrophage development will be examined and a search for the exact function of the receptor will be made. 3) The structural basis for the binding of carbohydrate to the receptor will be sought.

三组蛋白质的结构与功能关系 参与糖缀合物生物合成的方面，或 功能将被检查。 起作用的糖基转移酶 在糖缀合物中寡糖的生物合成中， 研究了几种方法。 1)λ中的cDNA表达文库 将用针对某些转移酶的抗体筛选GTL 1， 分离含有转移酶cDNA的克隆， 测序 2)某些转移酶的cDNA将被插入 转化到合适的表达载体中以获得足够的 用于结构/功能研究的转移酶或特异性地获得 通过位点特异性诱变修饰酶。 3)两 在粘蛋白寡糖生物合成中起作用的转移酶， 将被净化和表征。 的 两个转移酶形成核心序列GA 1 β 1，3Ga 1 NAc-O-， 丝氨酸/苏氨酸 用于润滑胃肠的粘蛋白类， 呼吸道和泌尿生殖道，并保护上皮细胞 从受伤和脱水将被隔离，并进一步 其特征如下。 1)猪的cDNA序列 将完成颌下腺cDNA的测序， 检查颌下粘蛋白的基因组DNA。 此外该 猪基因组中粘蛋白的基因数量将是 测定 2)粘蛋白抗体和合成多核苷酸 将使用基于颌下粘蛋白cDNA序列的方法， 确定其他猪组织是否含有颌下 粘蛋白样蛋白或mRNA。 它也将与使用确定 在这些探针中，人颌下无粘蛋白是否 结构上与猪颌下脱粘蛋白相关。 3)的 将确定寡糖在粘蛋白中的确切位置。 四、 来自其他猪组织的粘蛋白，如胃、小 肠和结肠将被分离并进行结构表征。 低聚糖巨噬细胞受体 研究，特别是只在大鼠和小鼠上发现的受体 枯否细胞。 1)大鼠基因组DNA序列测定 将测定枯否细胞。 2)的表达 受体在巨噬细胞发育过程中将进行检查， 将对受体的确切功能进行研究。 第三章 碳水化合物结合到 将寻求接收器。