STRUCTURE DETERMINATION OF ION CHANNEL PROTEINS

离子通道蛋白的结构测定

• 批准号: 8361661
• 负责人: BENOIT ROUX
• 金额: $ 2.19万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-04-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8361661
• 关键词: Anions; Antibiotic Resistance; Antibiotics; Binding; Brain; Cations; Cell Membrane Permeability; Cells; Characteristics; Charge; Complex; Crystallography; Drug Interactions; Funding; Glutamate Receptor; Gram-Negative Bacteria; Grant; Ion Channel; Ion Channel Protein; Ions; Ligand Binding Domain; Mediating; Membrane; Molecular; National Center for Research Resources; Neurotransmitters; Pathway interactions; Principal Investigator; Property; Proteins; Research; Research Infrastructure; Resources; Source; Specificity; Spinal Cord; Structure; Synapses; Time; United States National Institutes of Health; Work; antibiotic design; aqueous; computer studies; constriction; cost; dimer; electric field; extracellular; improved; insight; novel; porin; receptor; response; solute; structural biology

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. OmpF porins are cation-selective aqueous channels found in the outer membrane (OM) of Gram-negative bacteria. Their main function is to facilitate the translocation of small hydrophilic solutes across the OM. One unique feature of the OmpF porin is an extracellular loop that folds into the interior of the pore, creating the constriction zone. It has been suggested that charge asymmetry in the constriction zone could give rise to a strong electric field and aid in permeation of dipolar solutes. Computational studies have suggested an intriguing ion-conducting property of OmpF, in which cations and anions follow separated pathways. However, this feature has yet to be directly observed experimentally. We are, therefore, employing x-ray crystallography in order to study the cation and anion selective pathways along the OmpF pore. In addition to its unique ion conduction properties, the OmpF channel is also the main gateway that allows antibiotic molecules to permeate Gram-negative bacterial cells. With the emergence of antibiotic resistance by virtue of decreased membrane permeability, it is necessary to find ways to increase antibiotic translocation across OmpF porin. Therefore, in a second subproject, we have crystallized OmpF in complex with various antibiotics. For the first time, we are able to experimentally visualize protein-drug interactions at the molecular level. Therefore, results of this work will give insights into the design of antibiotics with improved diffusional characteristics and target specificity. The ionotropic glutamate receptor ion channels (iGluRs) mediate excitatory responses at the vast majority of synapses in the brain and spinal cord. The binding of neurotransmitter molecules to the ligand-binding domain (LBD) of these receptors drives the opening of the transmembrane pore. These receptors assemble as tetramers. However, previous crystal structures have revealed only dimer complexes. Therefore, in this third project, we have solved the crystal structure of a novel GluR2 LBD dimer-of-dimers.

该子项目是利用资源的众多研究子项目之一 由 NIH/NCRR 资助的中心拨款提供。子项目的主要支持 并且子项目的主要研究者可能是由其他来源提供的， 包括其他 NIH 来源。 子项目可能列出的总成本 代表子项目使用的中心基础设施的估计数量， NCRR 赠款不直接向子项目或子项目工作人员提供资金。 OmpF 孔蛋白是革兰氏阴性细菌外膜 (OM) 中发现的阳离子选择性水通道。 它们的主要功能是促进小的亲水性溶质穿过 OM 的易位。 OmpF 孔蛋白的一个独特特征是细胞外环折叠到孔内部，形成收缩区。 有人认为，缢缩区的电荷不对称性可能会产生强电场并有助于偶极溶质的渗透。 计算研究表明 OmpF 具有有趣的离子传导特性，其中阳离子和阴离子遵循不同的路径。 然而，这一特征尚未通过实验直接观察到。 因此，我们采用 X 射线晶体学来研究沿 OmpF 孔的阳离子和阴离子选择性路径。 除了其独特的离子传导特性外，OmpF通道也是抗生素分子渗透革兰氏阴性细菌细胞的主要门户。 随着膜通透性降低而出现抗生素耐药性，有必要找到增加抗生素跨 OmpF 孔蛋白易位的方法。 因此，在第二个子项目中，我们将 OmpF 与各种抗生素复合结晶。 我们第一次能够在分子水平上通过实验可视化蛋白质-药物相互作用。 因此，这项工作的结果将为设计具有改善的扩散特性和靶点特异性的抗生素提供见解。离子型谷氨酸受体离子通道 (iGluR) 介导大脑和脊髓绝大多数突触的兴奋反应。神经递质分子与这些受体的配体结合域（LBD）的结合驱动跨膜孔的打开。 这些受体组装成四聚体。 然而，以前的晶体结构仅揭示了二聚体复合物。 因此，在第三个项目中，我们解决了新型 GluR2 LBD 二聚体的晶体结构。