NMR STUDIES OF BIOLOGICAL MEMBRANES

生物膜的核磁共振研究

• 批准号: 6635941
• 负责人: JUDITH HERZFELD
• 金额: $ 25.85万
• 依托单位: BRANDEIS UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 1985
• 资助国家: 美国
• 起止时间: 1985-09-30 至 2004-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6635941
• 关键词: Halobacteriaceae; active sites; bacteriorhodopsins; bioimaging /biomedical imaging; cell membrane; chemical bond; chloride ion; chromophore; high energy particle; hydrogen ions; infrared spectrometry; interferometry; intermolecular interaction; ion transport; membrane activity; membrane structure; nuclear magnetic resonance spectroscopy; photochemistry; structural biology

Description: (Verbatim from the applicant's abstract) A distinctive feature of living cells is the facility with which they convert energy from one form to another. Central among these energy transactions is the active transport of ions across cell membranes. In recent years, significant progress has been made in elucidating the structures of various ion pumps. However, the molecular mechanisms by which vectorial ion motion is enforced remain unknown. Our long-term goal is to understand how proteins carry out this essential aspect of energy transduction. A particularly propitious system for this purpose is the light-driven proton pump, bacteriorhodopsin. Key steps in the photocycle have been identified and the structure of the resting state has been established by diffraction methods (with the exception of a few disordered, packing sensitive, or hydration sensitive regions). Our goal is to obtain a detailed picture of the active site of the molecule as it evolves through the critical steps of the photocycle. In particular, we are concerned with the structural changes that occur around the Schiff base of the retinal chromophore while it is deprotonated (i.e., during the M stage of the photocycle), because these changes prevent the proton that is released to the extracellular side from returning when the Schiff base reprotonates. Thus we are looking specifically for a switch in the connectivity of the deprotonated Schiff base between transport pathways on the two sides of the membrane and clues to the source of the irreversibility of this switch. Since our work was last funded, we have identified early and late M intermediates and learned how to trap them at levels suitable for solid state NMR studies. In addition, the last few years have seen a dramatic expansion of the range and power of solid state NMR techniques and we have developed procedures for new variations in the isotopic labeling of bacteriorhodopsin that will allow us to take advantage of the new spectroscopy. Bringing these elements together, we will characterize the spatial relationships and chemical exchange connectivities that define the active site and how they change in the critical photocycle intermediates.

描述：（逐字研究来自申请人的摘要） 活细胞是将能量从一种形式转化为 其他。这些能源交易中的中心是主动运输 跨细胞膜的离子。近年来，取得了重大进展 阐明各种离子泵的结构。但是，分子 实施矢量离子运动的机制尚不清楚。我们的 长期目标是了解蛋白质如何进行这一基本方面 能量转导。 为此目的的特别好的系统是轻驱动质子 泵，细菌紫红质。已经确定了光循环的关键步骤，并且 静止状态的结构已通过衍射方法建立 （除了几个无序，包装敏感或水合 敏感区域）。我们的目标是获取活动网站的详细图片 分子通过光圈的临界步骤演变而来的。在 特别是，我们关心周围发生的结构变化 视网膜发色团的席夫基部被去质子化时（即，在 光圈的M阶段），因为这些变化阻止了质子 当席夫基地时，被释放到细胞外侧 重体。因此，我们正在专门寻找连接性的开关 在两侧的两侧的运输途径之间的去质子化的席夫底座 该开关的不可逆性来源的膜和线索。 由于我们的工作是最后资助的，我们已经确定了早期和晚期 中级并学会了如何以适合固态的水平捕获它们 NMR研究。此外，最近几年已经看到了巨大的扩张 固态NMR技术的范围和力量，我们已经开发了 细菌淡淡蛋白质的同位素标记的新​​变化程序 这将使我们能够利用新的光谱法。带来这些 元素在一起，我们将表征空间关系和化学 交换连接定义活动站点及其如何更改的连接性 临界光循环中间体。