Structural Analysis of the ATP Synthase Membrane Domain

ATP 合酶膜结构域的结构分析

• 批准号: 6525897
• 负责人: MARK E. GIRVIN
• 金额: $ 30.33万
• 依托单位: ALBERT EINSTEIN COLLEGE OF MEDICINE
• 依托单位国家: 美国
• 财政年份: 1997
• 资助国家: 美国
• 起止时间: 1997-01-01 至 2005-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6525897
• 关键词: Escherichia coli; acidity /alkalinity; active sites; affinity chromatography; artificial membranes; chemical bond resonance; chimeric proteins; computer simulation; conformation; crystallization; dimer; high performance liquid chromatography; hydrogen transporting ATP synthase; mass spectrometry; model design /development; molecular dynamics; monomer; nuclear magnetic resonance spectroscopy; physical model; protein folding; protein purification; protein structure function; protonation; stable isotope double label; structural biology; thermodynamics

DESCRIPTION (provided by applicant): The F1F0 ATP synthase is responsible for synthesizing the vast majority of cellular AlP. Not surprisingly, deleterious mutations in genes of the ATP synthase lead to inherited disorders, especially of nerves and muscles. The enzyme consists of two subcomplexes. The water-soluble F1 contains the catalytic sites for AlP synthesis and hydrolysis. The transmembrane F0 is responsible for proton transport. Remarkable progress has been made in understanding the structural and mechanistic aspects of catalysis by F1. As is always the case with membrane proteins, progress with the Fo has been much slower. Fo comprises three types of subunits in an a1b2c10 stoichiometry. Proton translocation through Fo is hypothesized to occur at the interface between the a-and c-subunits, beginning with a half-channel in subunit-a, moving through the essential Asp6l of subunit-c, and concluding with a second half-channel in subunit-a. Recent structures of subunit-c monomers in both protonation state suggest that during proton translocation the C-terminal helix of subunit-c rotates against subunit-a as a small "gear," driving rotation of a ring of c-subunits relative to the static subunits, and ultimately leading to the catalytic conformational changes in F1. This hypothesis will be tested by: 1) solving the structure of subunit-c in its oligomeric form, 2) determining the active site configurations of the c-subunits during the steps of proton translocation, 3) identifying the proton translocation pathways in subunit-a, and 4) determining how access of the active site Asp61 residue of subunit-c is limited to one side of the membrane at a time. The sample conditions and NMR methodologies developed to accomplish these aims will have general application to studying membrane protein structure by NMR. Membrane proteins are responsible for transmembrane signaling, energy transduction, and ion and metabolite transport. These proteins are important in infectious disease, genetic disorders, and cancer. Despite their importance, and the need for structure to understand their function, few such structures are available.

描述（由申请人提供）：F1F0 ATP合酶负责 合成绝大多数细胞 AlP。毫不奇怪，有害 ATP合酶基因突变会导致遗传性疾病，尤其是 神经和肌肉。该酶由两个子复合物组成。这 水溶性 F1 含有 AlP 合成和水解的催化位点。 跨膜 F0 负责质子传输。显着进步 已经在了解结构和机械方面进行了 F1催化。与膜蛋白的情况一样，进展与 Fo 的速度要慢得多。 Fo 在 a1b2c10 中包含三种类型的亚基 化学计量。假设质子通过 Fo 的易位发生在 a 和 c 亚基之间的界面，以半通道开始 亚基-a，遍历亚基-c 的基本 Asp6l，最后得出 子单元 a 中的第二个半通道。亚基-c单体的最新结构 两种质子化状态表明，在质子易位过程中，C 端 亚基-c 的螺旋作为一个小“齿轮”相对于亚基-a 旋转，驱动 c 亚基环相对于静态亚基的旋转，以及 最终导致F1的催化构象变化。这 假设将通过以下方式进行检验： 1）求解其子单元-c 的结构 寡聚形式，2) 确定活性位点构型 质子易位步骤中的 c 亚基，3) 识别质子 亚基-a中的易位途径，以及4)确定如何访问 亚基-c 的活性位点 Asp61 残基仅限于膜的一侧 一次。 为实现这些目标而开发的样品条件和 NMR 方法 将普遍应用于通过核磁共振研究膜蛋白结构。 膜蛋白负责跨膜信号传导、能量传递 转导、离子和代谢物运输。这些蛋白质很重要 传染病、遗传性疾病和癌症。尽管它们很重要， 以及需要结构来理解其功能，很少有这样的结构 可用。