F1-ATPase Chemical-Mechanical Coupling Mechanisms

F1-ATP酶化学机械耦合机制

• 批准号: 8005216
• 负责人: WAYNE D FRASCH
• 金额: $ 18.02万
• 依托单位: ARIZONA STATE UNIVERSITY-TEMPE CAMPUS
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-12-31 至 2010-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8005216
• 关键词: ADP-AlF4; ATP Hydrolysis; ATP Synthesis Pathway; ATP phosphohydrolase; Affect; Affinity; Bacteria; Binding; Biological Assay; C10; Catalytic Domain; Cell membrane; Charge; Chemicals; Chloroplasts; Communication; Complex; Computer software; Coupling; Custom; Enzymes; Escherichia coli; Event; F1-ATPase; Fluorescence; Generations; Goals; Human; Hydrogen Bonding; Ions; Laboratories; Location; Measurement; Measures; Mechanics; Membrane; Metals; Microscope; Mitochondria; Modeling; Molecular Motors; Motion; Movement; Mutation; Noise; Nucleotides; Protons; Reaction; Research Personnel; Rotation; Signal Transduction; Sodium Chloride; Structure; Testing; Thermodynamics; Time; Torque; Tyrosine; Work; Writing; clinically relevant; driving force; innovation; mutant; programs; single molecule

The F1Fo ATP synthase is a multisubunit, membrane-bound enzyme that catalyzes the synthesis of the majority of cellular ATP.Mutations in several subunits in the human mitochondrial enzyme are of important clinical relevance. The enzyme works as a molecular motor that drives the rotary motion of some of the subunits. The enzymes from mitochondria, chloroplasts and the plasma membranes of bacteria are closely similar. In E. coli, F1 is composed of an (ap)3 ring that surrounds the y subunit, and also contains 5 and e subunits. The membrane embedded Fo is composed of a c10 subunit ring to which is attached the a subunit and two b subunits. During ATP synthesis, the movement of protons through Fo drives the rotation of the c10 subunit ring to which the y and esubunits are attached that forces sequential conformational changes in the (cc|3)3 ring and results in the synthesis of ATP from each of the three (ap)3 heterodimers. The hydrolysis of ATP can also drive the rotation of the y subunit in the opposite direction. When F1 molecules are attached to a microscope cover slip, and a probe that can be observed under a microscope is attached to the y subunit, single molecules can be observed to rotate upon addition of ATP. Our long term objective is to elucidate the mechanism of ATPase-driven rotation of the y subunit. We propose the induction mechanism as a working hypothesis for rotation that will be critically examined. This hypothesis posits that Coulombic potential originating from residues that form hydrogen bonds and salt bridges between the (a(3)3 ring and the y subunit contribute to the generation of y subunit torque. We will test this hypothesis by accomplishing the following aims. (1) The contribution of p Catch Loop-y Subunit interactions to y subunit rotation will be examined by measuring the effects of mutations that eliminate these interactions on the rate of y subunit rotation measured using an innovative single molecule assay that we developed for this purpose. (2) The contribution of other known (a(J)3 ring-y subunit interactions to y subunit rotation will be determined that include contacts at the y subunit N and C termini.(3) The contribution to rotation of (ap)3 ring residues that do not interact directly with the y subunit will be examined including those that communicate between the y subunit and the catalytic sites. (4) The contribution to y subunit rotation of (ap)3 ring-y subunit interactions that have not been identified in known crystal structures of F1 but are identified by tracking the rotational path of charged and polar y subunit residues around the inside of the (ap)3 ring.

F1FO ATP合酶是一种多育膜结合的酶，可催化大多数的合成 细胞ATP。人线粒体酶中几个亚基的过多是重要的临床相关性。这 酶是一种分子运动，可驱动某些亚基的旋转运动。来自 线粒体，叶绿体和细菌的质膜非常相似。在大肠杆菌中，F1由 （AP）3环围绕Y亚基，还包含5个和E亚基。膜嵌入的FO组成 附着A亚基和两个B亚基的C10亚基环。在ATP合成过程中， 质子通过FO驱动C10亚基环的旋转 （CC | 3）3环的顺序构象变化，并导致三个（AP）3中的每个 异二聚体。 ATP的水解还可以驱动Y亚基的旋转，以相反的方向驱动。当F1时 分子连接到显微镜盖盖上，并且可以在显微镜下观察到的探针附着在 Y亚基，可以观察到单分子在添加ATP时旋转。我们的长期目标是阐明 Y亚基ATPase驱动旋转的机理。我们提出诱导机制作为一种工作 旋转的假设将进行严格检查。该假设认为库仑的潜力源自 在（a（a（3）3环和Y亚基）之间形成氢键和盐桥的残基有助于 Y亚基扭矩的产生。我们将通过实现以下目标来检验这一假设。 （1）P的贡献 通过测量突变的影响，将检查与Y亚基旋转的捕获环-Y亚基相互作用 消除使用创新的单分子测定法测量的Y亚基旋转速率的相互作用 我们为此目的开发了。 （2）其他已知的其他已知（a（j）3环亚基相互作用对y亚基旋转的贡献 将确定包括在y亚基N和C末端的触点。（3）（3）（AP）3环的旋转的贡献 将检查不直接与Y亚基相互作用的残留物，包括在y之间通信的残留物 亚基和催化位点。 （4）对（AP）3个环形亚基相互作用的y亚基旋转的贡献 已在F1的已知晶体结构中鉴定出来，但通过跟踪带电和极性Y的旋转路径来识别 （AP）3环内部周围的亚基残基。

项目成果

Single-molecule detection of DNA via sequence-specific links between F1-ATPase motors and gold nanorod sensors.

• DOI: 10.1039/b716744j
• 发表时间: 2008-02
• 期刊: Lab on a chip
• 影响因子: 6.1
• 作者: J. York;D. Spetzler;Fusheng Xiong;W. Frasch

Structural Asymmetry and Kinetic Limping of Single Rotary F-ATP Synthases.

单旋转 F-ATP 合成酶的结构不对称性和动力学跛行。

• DOI: 10.3390/molecules24030504
• 发表时间: 2019
• 期刊: Molecules (Basel, Switzerland)
• 作者: Sielaff,Hendrik;Yanagisawa,Seiga;Frasch,WayneD;Junge,Wolfgang;Börsch,Michael
• 通讯作者: Börsch,Michael

Interactions between beta D372 and gamma subunit N-terminus residues gamma K9 and gamma S12 are important to catalytic activity catalyzed by Escherichia coli F1F0-ATP synthase.

β D372 和 γ 亚基 N 末端残基 γ K9 和 γ S12 之间的相互作用对于大肠杆菌 F1F0-ATP 合酶的催化活性很重要。

• DOI: 10.1021/bi047293j
• 发表时间: 2005
• 期刊: Biochemistry.
• 作者: Lowry,DavidS;Frasch,WayneD
• 通讯作者: Frasch,WayneD

Interactions among gamma R268, gamma Q269, and the beta subunit catch loop of Escherichia coli F1-ATPase are important for catalytic activity.

γ R268、γ Q269 和大肠杆菌 F1-ATP 酶的 β 亚基捕获环之间的相互作用对于催化活性非常重要。

• DOI: 10.1074/jbc.m309948200
• 发表时间: 2003
• 期刊: The Journal of biological chemistry
• 作者: Greene,MatthewD;Frasch,WayneD
• 通讯作者: Frasch,WayneD

Abundance of Escherichia coli F1-ATPase molecules observed to rotate via single-molecule microscopy with gold nanorod probes.

通过使用金纳米棒探针的单分子显微镜观察到大量的大肠杆菌 F1-ATPase 分子旋转。

• DOI: 10.1007/s10863-007-9114-x
• 发表时间: 2007
• 期刊: Journal of bioenergetics and biomembranes
• 影响因子: 3
• 作者: York,Justin;Spetzler,David;Hornung,Tassilo;Ishmukhametov,Robert;Martin,James;Frasch,WayneD
• 通讯作者: Frasch,WayneD