GLUCOSYLTRANSFERASE IN MAMMARY GLAND

乳腺中的葡萄糖基转移酶

• 批准号: 3303870
• 负责人: INDER K VIJAY
• 金额: $ 7.33万
• 依托单位: UNIV OF MARYLAND, COLLEGE PARK
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1993-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3303870
• 关键词: affinity chromatography; affinity labeling; antibody formation; dolichol; enzyme structure; glucose metabolism; glycoprotein biosynthesis; glycosyltransferase; hormone regulation /control mechanism; immunoprecipitation; laboratory rabbit; laboratory rat; lactation; mammary gland; molecular cloning; protein purification; transposon /insertion element; western blottings

The biosynthesis of N-linked glycoproteins involves a step wise, dolichol- linked assembly of a tetrasaccharide precursor, Glc3Man9GlcNAc2 and its en bloc transfer to the nascent peptide followed by co- and post-translational modifications of the oligosaccharyl moiety. The donor for the glycosyl residues in the precursor is Glc-P-Dol; its biosynthesis is catalyzed by UDP-Glc:Dol-P glucosyltransferase. It has been shown that glucosylation of the precursor signals its transfer to the acceptor polypeptides. One may hypothesize that the action of glucosyltransferase represents a key step for the overall regulation of N-glycosylation. The mammary gland possesses some unique features for studying the biosynthesis and regulation of glycoproteins. It is under intense hormonal modulation for its growth and development as it repeatedly cycles between dormancy -> gestation -> lactation -> regression -> dormancy throughout the reproductive life of the female. Further, the lactating gland synthesizes and secretes massive amounts of only a limited number of well characterized glycoproteins into milk. Previous studies from our laboratory have elucidated the outlines of the dolichol cycle and the stereochemical structures of the intermediates of the cycle in the mammary gland. The first enzyme, i.e., UDP-GlcNAc:Dol- P GlcNAc-1-P-transferase and processing-specific glucosidases I and II have been purified. The proposed program will purify, characterize and clone the glucosyltransferase. Innovative methodologies, based on the synthesis of novel analogs of the enzyme substrate will be incorporated in the purification protocol. The enzyme cDNA will be cloned in the expression vector lambdaZAP-II. IN studies outside the scope of this work, molecular probes for glucosyltransferase will be employed to investigate the gland. The recent pioneering studies on the expression of glycosylated and functional glycoproteins tPA, alpha-antitrypsin and blood clotting factor IX in the milk of transgenic animals illustrate the power of the mammary gland as a bioreactor of immense potential for biomedical and agrochemical purposes. Glycoproteins participate in a wide spectrum of biological functions and recognition phenomena. Pathological states including malignancy, atherosclerosis and many autosomal recessive disorders are accompanied with marked alterations in cellular glycoprotein metabolism. A glycoprotein-glycoprotein interaction between gp120 of HIV and CD4 receptor on the lymphocyte provides the entry port for the introduction of viral genetic material into the host leading to AIDS.

N-连接糖蛋白的生物合成涉及一个逐步的多萜醇- 四糖前体Glc 3 Man 9 GlcNAc 2及其酶的连接组装 阻断转移到新生肽，然后进行共翻译和翻译后 寡糖基部分的修饰。 糖基的供体 前体中的残基是Glc-P-Dol;其生物合成由 UDP-Glc：Dol-P葡糖基转移酶。 已经表明， 前体发出信号将其转移到受体多肽。 人们可以 假设葡糖基转移酶的作用代表了一个关键步骤， 用于N-糖基化的整体调节。 乳腺具有 研究生物合成和调控的一些独特的功能 糖蛋白 它的生长受到强烈的荷尔蒙调节， 在休眠->妊娠-> 哺乳->退化->休眠在整个生殖生命的 女性 此外，泌乳腺合成和分泌大量的 量的仅有限数量的充分表征的糖蛋白转化为 牛奶 我们实验室以前的研究已经阐明了 多萜醇环和中间体的立体化学结构 在乳腺的周期。 第一种酶，UDP-GlcNAc：Dol- P GlcNAc-1-P-转移酶和加工特异性葡糖苷酶I和II具有 被净化了 拟议的计划将纯化，表征和克隆 葡糖基转移酶 在综合报告基础上的创新方法 酶底物的新型类似物将被掺入到 纯化方案。 将在表达载体中克隆酶cDNA。 载体peptidaZAP-II。 在这项工作范围之外的研究中，分子 葡糖基转移酶的探针将用于研究腺体。 近年来，关于糖基化和 功能性糖蛋白tPA、α-抗胰蛋白酶和凝血因子 九在转基因动物的牛奶说明了权力的乳腺癌 腺体作为生物医学和农业化学的巨大潜力的生物反应器 目的 糖蛋白参与多种生物学过程， 功能和识别现象。 病理状态包括 恶性肿瘤、动脉粥样硬化和许多常染色体隐性遗传疾病是 伴有细胞糖蛋白代谢的显著改变。 一 HIV gp 120与CD 4受体糖蛋白相互作用 淋巴细胞上的淋巴细胞为病毒的进入提供了入口 遗传物质进入宿主体内导致艾滋病