Influence of lipids on membrane protein structure and function

脂质对膜蛋白结构和功能的影响

• 批准号: 7924968
• 负责人: Tamir Gonen
• 金额: $ 26.18万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-30 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7924968
• 关键词: Address; Adherens Junction; Affect; Binding; Biological; Biological Assay; Biological Models; C-terminal; Calcium; Calmodulin; Cell physiology; Cells; Chemistry; Collaborations; Complex; Crystalline Lens; Crystallization; Cytoplasm; DNA Sequence Rearrangement; Electrophysiology (science); Environment; Face; Goals; Housing; Intercellular Junctions; Lateral; Lens Fiber; Ligands; Lipid Bilayers; Lipids; Location; MIP gene; Mediating; Membrane; Membrane Biology; Membrane Lipids; Membrane Proteins; Oocytes; Permeability; Physiological; Proteins; Recombinants; Research Personnel; Resolution; Role; Signal Transduction; Structure; Structure-Activity Relationship; System; Tail; Universities; Water; cell type; electron crystallography; extracellular; fiber cell; insight; lens; oxidation; periplasm; professor; programs; protein structure function; reconstitution; research study; response; saturated fat; tool; two-dimensional; water channel

DESCRIPTION (provided by applicant): In membranes, proteins and lipids form an integrated system to support essential cell functions. The structure of membrane proteins embedded in native lipid bilayers can be studied by electron crystallography. Aquaporin 0 (AQPO) is the major protein constituent in lens fiber cells where it serves a dual role: creating a pore for water and forming adhesive junctions. Using electron crystallography, we have recently determined the structure of AQPO to an unprecedented resolution of 1.9A, revealing the precise structure of a shell of ordered lipids surrounding the AQPO tetramer. AQPO therefore provides the first eukaryotic model system in which it is possible to study, in the very same membrane, the structural and functional interdependence of protein and lipids. To do this, we will form single-layered two-dimensional crystals of AQPO in lipid bilayers using native or unnatural lipids and in the presence or absence of calmodulin (CaM) which binds to the C- terminal domain of AQPO. We will then correlate the structure (from electron crystallography) and function (from electrophysiology) of the AQPO water channel as it opens and closes in response to pH, divalent calcium, and other ligands. Finally, we will also determine the structural rearrangements required for pore closure when two AQPO tetramers on separate bilayers come together to form intercellular junctions. Although this proposal addresses specific questions regarding AQPO water channel function, many of the resulting insights should be generally applicable to other channels and membrane proteins.

描述（由申请人提供）：在膜中，蛋白质和脂质形成了支持基本细胞功能的集成系统。可以通过电子晶体学研究嵌入天然脂质双层中的膜蛋白的结构。 Aquaporin 0（AQPO）是透镜纤维细胞中的主要蛋白质成分，它具有双重作用：为水创建孔和形成粘合剂连接。使用电子晶体学，我们最近确定了AQPO的结构至1.9a的前所未有的分辨率，揭示了AQPO四聚体周围有序脂质的壳的精确结构。因此，AQPO提供了第一个真核模型系统，可以在相同的膜上研究蛋白质和脂质的结构和功能相互依赖性。为此，我们将使用天然或非天然脂质在脂质双层中形成AQPO的单层二维晶体，​​并在存在或不存在与AQPO的C-末端结构域结合的存在或不存在的情况下。然后，我们将在AQPO水通道的结构（来自电子晶体学）和功能（来自电生理学）相关联，因为它响应于pH，二价钙和其他配体而打开并关闭。最后，当两个单独的双层中的两个AQPO四聚体组合在一起形成细胞间连接时，我们还将确定孔隙闭合所需的结构重排。尽管该提案解决了有关AQPO水通道功能的具体问题，但许多结果的见解通常应适用于其他通道和膜蛋白。