MASS SPECTROMETRIC STUDIES OF INTEGRAL MEMBRANE PROTEINS & ION CHANNELS

完整膜蛋白的质谱研究

• 批准号: 8169097
• 负责人: RODERICK MACKINNON
• 金额: $ 0.47万
• 依托单位: ROCKEFELLER UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-03-01 至 2011-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8169097
• 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; 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.

该副本是利用众多研究子项目之一 由NIH/NCRR资助的中心赠款提供的资源。子弹和 调查员（PI）可能已经从其他NIH来源获得了主要资金， 因此可以在其他清晰的条目中代表。列出的机构是 对于中心，这不一定是调查员的机构。 我们正在努力开发质谱法的实用性，以帮助使用衍射方法在原子分辨率下定义积分膜蛋白和（尤其是离子通道）的定义。这项工作导致了许多出版物。 Doyle，J.M。Cabral，R.A。 Pfuetzner，A。Kuo，J.M。Gulbis，S.L。科恩（B.T.） Chait，R。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。 Cohen，A。Kuo，A。Lee，B.T。 Chait“原核生物和真核钾通道中的结构保护”科学280（1998）106-109。 J.H. Morais Cabral，A。Lee，S。Cohen，B.T。 Chait，M。Li，R。Mackinnon“ HERG钾通道氨基末端结构域的结构：PAS域结构家族的定义” Cell 95（1998）649-655。 J.M. Gulbis，M。Zhou，S。Mann，R。Mackinnon“胞质的结构（电压依赖性K+通道的亚基T1组装” Science 289（2000）123-127。M.Cadene，B.T。Chait” B.T. Chait”稳健的清洁剂，用于质谱分析的强大清洁剂，对整合性分析70（2000）。 5655-5658。阴离子的选择性”自然415（2002）287-94。Y.Jiang，A。Lee，J。Chen，M。Cadene，B。T. Chait，R。Mackinnon，R。Mackinnon，“钙门控钾通道的晶体结构和机制”自然417（2002）（2002）515-522。515-522。515-522。Y.Jiang，A。Lee，A。Lee，J.Chen，M。Chait，R。CADN，R。CADN，R。CADN，M。CADN，R。CADN，R。CADN，R.钾通道的孔构象”自然417（2002）523-526。Y.Jiang，A。Lee，J。Chen，V。Ruta，M。Cadene，B.T。 Chait＆Roderick Mackinnon“电压依赖性K+通道的X射线结构”自然423（2003）33-41。最近的工作涉及KIR3.1 K（+）通道，该通道通过G蛋白和脂质信号通路参与心率控制和神经元兴奋性。通过用原核生物KIR通道的孔代替四分之三的跨膜孔，使大肠杆菌的表达实现了，从而使kir3.1起源的细胞质孔隙和膜界面区域。在2.2 A下确定了两个结构。选择性滤波器在测量误差（R.M.S.D. <0.2 A）内与Lividans K（+）通道相同，这表明K（+）选择性需要对三维结构进行极大的保护。多个k（+）离子驻留在孔内，有助于解释电压依赖性的Mg（2+）和多胺阻滞和强烈的整流。在内螺旋束和细胞质孔的顶点处的两个限制可能充当门：在一种结构中，顶点是打开的，另一个结构是封闭的。顶点的门控是由细胞质孔亚基的刚体运动介导的。磷脂酰肌醇4,5-二磷酸互动残基提出了一种可能的机制，信号传导脂质调节细胞质孔。