MASS SPECTROMETRIC STUDIES OF INTEGRAL MEMBRANE PROTEINS & ION CHANNELS

完整膜蛋白的质谱研究

• 批准号: 7954048
• 负责人: RODERICK MACKINNON
• 金额: $ 0.71万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-03-01 至 2010-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7954048
• 关键词: Anions; Anus; Biophysics; Calcium; Cells; Chloride Channels; Computer Retrieval of Information on Scientific Projects Database; Cytoplasmic Structures; Detergents; Escherichia coli; Family; Funding; GTP-Binding Proteins; Grant; Heart Rate; Institution; Integral Membrane Protein; Ion Channel; Ions; Left; Lipids; Mass Spectrum Analysis; Measurement; Mediating; Membrane; Methods; Molecular; Molecular Conformation; Movement; Nature; Phosphatidylinositols; Polyamines; Potassium Channel; Publications; Research; Research Personnel; Resolution; Resources; Roentgen Rays; Ruta; Science; Signal Pathway; Signal Transduction; Source; Streptomyces lividans; Structure; United States National Institutes of Health; Work; base; constriction; interfacial; macromolecule; neuronal excitability; protein structure; three dimensional structure; tool; voltage

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. We are making a major effort to develop the utility of mass spectrometry to assist in the definition of integral membrane proteins and (especially ion channels) at atomic resolutions using diffraction methods. A number of publications have resulted from this work D.A. Doyle, J.M. Cabral, R.A. Pfuetzner, A. Kuo, J.M. Gulbis, S.L. Cohen, B.T. Chait, R. MacKinnon, "The Structure of the Potassium Channel: Molecular Basis of K+ Conduction and Selectivity" Science 280, (1998) 69-77. R. MacKinnon, S.L. Cohen, A. Kuo, A. Lee, B.T. Chait "Structural Conservation in Prokaryotic and Eukaryotic Potassium Channels" Science 280 (1998) 106-109. J.H. Morais Cabral, A. Lee, S. Cohen, B.T. Chait, M. Li, R. MacKinnon "Structure of the HERG potassium channel amino-terminal domain: definition of a structural family of PAS domains" Cell 95 (1998) 649-655. J.M. Gulbis, M. Zhou, S. Mann, R. MacKinnon "Structure of the Cytoplasmic ( Subunit-T1 Assembly of Voltage-Dependent K+ Channels" Science 289(2000) 123-127. M. Cadene, B.T. Chait "A Robust Detergent-Friendly for Mass Spectrometric Analysis of Integral Membrane Proteins" Anal. Chem. 72 (2000) 5655-5658. S.L. Cohen, B.T. Chait "Mass Spectrometry as a tool for Studying Protein Structure" Ann. Reviews of Biophysics and Biomolecular Structure 30 (2001) 67-85. R. Dutzler, ER Campbell, M. Cadene, B.T. Chait, R.MacKinnon "X-ray structure of a ClC chloride channel at 3.0 A reveals the molecular basis of anion selectivity" Nature 415 (2002) 287-94. Y. Jiang, A. Lee, J. Chen, M. Cadene, B. T. Chait, R. MacKinnon "Crystal structure and mechanism of a calcium-gated potassium channel" Nature 417 (2002) 515-522. Y. Jiang, A. Lee, J. Chen, M. Cadene, B. T. Chait, R. MacKinnon "The open pore conformation of potassium channels" Nature 417 (2002) 523-526. Y. Jiang, A. Lee, J. Chen, V. Ruta, M. Cadene, B.T. Chait & Roderick MacKinnon "X-ray structure of a voltage-dependent K+ channel" Nature 423 (2003) 33-41. The most recent work involves the Kir3.1 K(+) channel, which participates in heart rate control and neuronal excitability through G-protein and lipid signaling pathways. Expression in Escherichia coli has been achieved by replacing three fourths of the transmembrane pore with the pore of a prokaryotic Kir channel, leaving the cytoplasmic pore and membrane interfacial regions of Kir3.1 origin. Two structures were determined at 2.2 A. The selectivity filter is identical to the Streptomyces lividans K(+) channel within error of measurement (r.m.s.d.<0.2 A), suggesting that K(+) selectivity requires extreme conservation of three-dimensional structure. Multiple K(+) ions reside within the pore and help to explain voltage-dependent Mg(2+) and polyamine blockade and strong rectification. Two constrictions, at the inner helix bundle and at the apex of the cytoplasmic pore, may function as gates: in one structure the apex is open and in the other, it is closed. Gating of the apex is mediated by rigid-body movements of the cytoplasmic pore subunits. Phosphatidylinositol 4,5-biphosphate-interacting residues suggest a possible mechanism by which the signaling lipid regulates the cytoplasmic pore.

这个子项目是许多研究子项目中的一个 由NIH/NCRR资助的中心赠款提供的资源。子项目和 研究者（PI）可能从另一个NIH来源获得了主要资金， 因此可以在其他CRISP条目中表示。所列机构为 研究中心，而研究中心不一定是研究者所在的机构。 我们正在作出重大努力，以开发质谱仪的效用，以协助定义的完整的膜蛋白和（特别是离子通道）在原子分辨率使用衍射方法。D.A. Doyle，J.M. Cabral，R.A. Pfuetzner，A.郭，J.M.古尔比斯公司科恩，B.T.柴特河MacKinnon，“钾通道的结构：K+传导和选择性的分子基础”Science 280，（1998）69-77。 R. MacKinnon，S.L.科恩，A. Kuo，黄毛菊A. Lee，B.T. Chait，“Proteins and Eukaryotic Potassium Channels”，Science 280（1998）106-109。 J.H. Morais Cabral，A.李，S。 科恩，B.T. Chait，M.利河，巴西-地MacKinnon“Structure of the HERG potassium channel amino-terminal domain：definition of a structural family of PAS domains”Cell 95（1998）649-655. J.M. Gulbis，M. Zhou，S.曼河MacKinnon“电压依赖性K+通道的细胞质（亚单位-T1组装）的结构”Science 289（2000）123-127。 M. Cadene，BT Chait“A Robust Detergent-Friendly for Mass Spectrometric Analysis of Integral Membrane Proteins”Anal. 72（2000）5655-5658。 S.L.科恩，B.T.质谱分析法作为研究蛋白质结构的工具>，生物物理学与生物分子结构回顾，30（2001）67-85。R. Dutzler，ER坎贝尔，M. Cadene，BT Chait，R.MacKinnon“X-ray structure of a C1C chloride channel at 3.0 A reveals the molecular basis of anion selectivity”Nature 415（2002）287-94. Y. Jiang，中国茶条A. Lee，J. Chen，M. Cadene，B. T.柴特河MacKinnon“Crystal structure and mechanism of a calcium-gated potassium channel”Nature 417（2002）515-522. Y. Jiang，中国茶条A. Lee，J. Chen，M. Cadene，B. T.柴特河MacKinnon“钾通道的开放孔构象”Nature 417（2002）523-526。Y. Jiang，中国茶条A. Lee，J. Chen，V. Ruta，M. Cadene，BT Chait & Roderick MacKinnon“电压依赖性K+通道的X射线结构”Nature 423（2003）33-41。最近的工作涉及Kir3.1 K（+）通道，其通过G蛋白和脂质信号通路参与心率控制和神经元兴奋性。通过用原核Kir通道的孔替换四分之三的跨膜孔，留下Kir3.1起源的细胞质孔和膜界面区域，已经实现了在大肠杆菌中的表达。在2.2A下测定了两种结构。选择性过滤器与变铅青链霉菌K（+）通道在测量误差（r.m.s.d.）范围内相同。<0.2 A），表明K（+）选择性需要三维结构的极端保守性。多个K（+）离子驻留在孔内，有助于解释电压依赖性Mg（2+）和多胺阻断和强整流。两个收缩，在内部螺旋束和在细胞质孔的顶点，可以作为门：在一个结构的顶点是开放的，在其他的，它是关闭的。顶端的门控由细胞质孔亚单位的刚体运动介导。磷脂酰肌醇4，5-二磷酸相互作用的残基表明一种可能的机制，信号脂质调节细胞质孔。