CHARACTERIZATION OF GLUCOSE TRANSPORTER FUNCTION

葡萄糖转运蛋白功能的表征

• 批准号: 3225033
• 负责人: CHAN Y. JUNG
• 金额: $ 10.42万
• 依托单位: STATE UNIVERSITY OF NEW YORK AT BUFFALO
• 依托单位国家: 美国
• 财政年份: 1978
• 资助国家: 美国
• 起止时间: 1978-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3225033
• 关键词: affinity labeling; chemical binding; chemical structure function; conformation; crosslink; diabetes mellitus; glucose transport; high performance liquid chromatography; human tissue; insulin sensitivity /resistance; laboratory mouse; laboratory rabbit; mass spectrometry; membrane proteins; molecular pathology; obesity; protein sequence; protein structure function; transport proteins

The long term goal of this proposal is to identify the molecular defect(s) underlying the insulin-resistance seen in human diabetes and obesity. More immediate goals are to understand the molecular mechanisms how the facilitative glucose transporters, a family in intrinsic transmembrane proteins, catalyze translocation of selected sugar molecules. We will use purified human erythrocyte glucose transporter (HEGT) and study its tertiar structure, particularly that of putative glucose channel. A hydropathy analysis of the amino acid sequence of this protein predicts that it contains a transmembrane domain made of twelve transmembrane segments, five of which are amphipathic and may line the purported aqueous channel. It also contains three intracellular and one extracellular nonmembrane domains We use this prediction as a working model in determining the channel structure. We will determine which of these twelve transmembrane segments are positioned in contact with the lipid bilayer, and which the aqueous channel. We will first establish the basic chemical methodology required for the studies. This will include separation and identification of the twelve transmembrane segments. To determine the native tertiary structure of these transmembrane segments. We will chemically crosslink and identify neighboring transmembrane segments, and identify the channel-forming segments and nonchannel-forming segments by measuring the accessibility of segments to transportable, covalently active glucose analogs (4ADG and 6ADG and lipophilic covalent probes, respectively, Based on these data, we will deduce the overall structure of the transmembrane domain. There are strong indication for the importance of intrinsic activity- regulation of glucose transporter function in glucose homeostasis in normal and diseased states. Understanding of this regulation of the intrinsic activity at the molecular level would be greatly assisted once we understan the tertiary structure of the glucose channel as proposed here.

该提案的长期目标是识别分子缺陷 人类糖尿病和肥胖症中的胰岛素抵抗是其背后的原因。 更多的 近期目标是了解分子机制 促进葡萄糖转运蛋白，内在跨膜家族 蛋白质，催化选定糖分子的易位。 我们将使用 纯化人红细胞葡萄糖转运蛋白（HEGT）并研究其三级结构 结构，特别是假定的葡萄糖通道的结构。 水病 该蛋白质的氨基酸序列分析预测它 包含一个由十二个跨膜片段组成的跨膜结构域，五个 其中是两亲性的并且可能排列在所谓的水通道中。 它 还包含三个细胞内和一个细胞外非膜结构域 我们使用这个预测作为确定通道的工作模型 结构。 我们将确定这十二个跨膜片段中的哪一个 位于与脂质双层接触的位置，并且水性 渠道。 我们将首先建立所需的基本化学方法 为了学习。 这将包括分离和识别 十二个跨膜节段。 确定天然三级结构 这些跨膜片段。 我们将进行化学交联并鉴定 相邻的跨膜片段，并识别通道形成 段和非通道形成段通过测量可访问性 片段为可运输的、共价活性的葡萄糖类似物（4ADG 和 6ADG 和亲脂性共价探针，基于这些数据，我们将 推导出跨膜结构域的整体结构。 有强烈的迹象表明内在活动的重要性 - 正常人葡萄糖稳态中葡萄糖转运蛋白功能的调节 和患病状态。 对这种内在规律的理解 一旦我们理解了，分子水平上的活动将会得到极大的帮助 此处提出的葡萄糖通道的三级结构。