STRUCTURE AND MECHANISM OF THE E COLI MANNITOL PERMEASE

大肠杆菌甘露醇渗透酶的结构和机制

• 批准号: 3275531
• 负责人: GARY R JACOBSON
• 金额: $ 14.16万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 1980
• 资助国家: 美国
• 起止时间: 1980-07-01 至 1994-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3275531
• 关键词: Escherichia coli; active transport; autoradiography; bacterial proteins; carbohydrate transport; conformation; cytoplasm; enzyme mechanism; genetic manipulation; mannitol; membrane permeability; membrane proteins; mutant; phospholipids; phosphorylation; protein sequence; protein structure function; pyruvate kinase; radiotracer; stereochemistry; transport proteins

Our laboratory is actively investigating the structure and mechanism of the enzyme permease responsible for the tightly coupled transport and phosphorylation of the hexito: D-mannitol in Escherichia coli. This protein is an integral membrane component of the bacterial phosphotransferase system (PTS) as well as serving as the primary chemotactic receptor for D-mannitol in E. coli. For the past 8 years, its structure and its catalytic mechanism have been studied in several laboratories, including our own. The broad objective of these studies is to determine in as much molecular detail as possible the mechanism by which the mannitol permease carries out its receptor and transport function. This is not completely known for any integral membrane transport protein, and our results should therefore add fundamental knowledge to the study of membrane protein structure and function. Since all cells rely on proper functioning of integral membrane proteins for interaction with their environment, and some diseased cells may lack one or more of theses functions, the study of these fundamental processes is essential in understanding the principles of growth and metabolism in both normal and diseased cells. In the proposed project continuation our aims are to: 1) continue our studies on the intramembrane structure of the membrane-bound domain of the mannitol permease; 2) study the structure of the cytoplasmic domain of the protein; 3) continue our studies on the mechanism of transport and phosphorylation catalyzed by this protein; 4) study structure-function relationships in this protein using in vitro mutagenesis techniques; and 5) isolate and characterize mutants in specific functions of the mannitol permease. A combination of biochemical, biophysical, molecular biological and genetic techniques will be used in these studies. Successful completion of this work will lead to significant progress toward our overall goal of understanding the molecular basis of transport in this system.

我们实验室正在积极研究其结构和机制 负责紧密耦合运输的酶通透性和 大肠杆菌中己糖醇的磷酸化：D-甘露醇。 这 蛋白质是细菌膜的组成部分 磷酸转移酶系统（PTS）以及作为主要 大肠杆菌中 D-甘露醇的趋化受体。 过去8年里， 其结构和催化机制已被国内外学者研究 实验室，包括我们自己的实验室。 这些研究的总体目标是 尽可能详细地确定其机制 甘露醇渗透酶执行其受体和转运 功能。 对于任何整体膜来说，这一点尚不完全清楚 运输蛋白质，因此我们的结果应该增加基础 研究膜蛋白结构和功能的知识。 自从 所有细胞都依赖于整合膜蛋白的正常功能 与环境的相互作用，一些患病细胞可能缺乏一种 或更多这些功能，对这些基本过程的研究是 对于理解生长和新陈代谢的原理至关重要 正常细胞和患病细胞。 在拟议的项目延续中，我们 目标是：1）继续我们对膜内结构的研究 甘露醇渗透酶的膜结合结构域； 2）研究 蛋白质胞质结构域的结构； 3）继续我们的 转运及磷酸化催化机制研究 这种蛋白质； 4) 研究该蛋白质的结构-功能关系 使用体外诱变技术； 5) 分离和表征 甘露醇渗透酶特定功能的突变体。 的组合 生物化学、生物物理、分子生物学和遗传技术 将在这些研究中使用。 这项工作的顺利完成将 导致我们在理解的总体目标方面取得重大进展 该系统中运输的分子基础。