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.

这个子项目是利用资源的许多研究子项目之一。 由NIH/NCRR资助的中心拨款提供。对子项目的主要支持 子项目的首席调查员可能是由其他来源提供的， 包括美国国立卫生研究院的其他来源。为子项目列出的总成本可能 表示该子项目使用的中心基础设施的估计数量， 不是由NCRR赠款提供给次级项目或次级项目工作人员的直接资金。 OmpF孔蛋白是一种阳离子选择性的水通道，存在于革兰氏阴性菌的外膜上。它们的主要功能是促进小的亲水性溶质在OM中的转移。OmpF孔蛋白的一个独特特征是一个细胞外环，它折叠到毛孔内部，形成收缩带。结果表明，收缩带中的电荷不对称可以产生较强的电场，有助于偶极溶质的渗透。计算研究表明，OmpF具有耐人寻味的离子导电特性，其中阳离子和阴离子遵循不同的路径。然而，这一特征还没有在实验上直接观察到。因此，我们正在使用X射线结晶学来研究沿OmpF孔的阳离子和阴离子的选择路径。 除了其独特的离子传导特性外，OmpF通道也是让抗生素分子渗透到革兰氏阴性细菌细胞的主要通道。随着由于膜通透性降低而产生的抗生素耐药性的出现，有必要寻找增加抗生素跨OmpF孔蛋白易位的方法。因此，在第二个子项目中，我们已经结晶了OmpF与各种抗生素的复合体。第一次，我们能够在分子水平上实验可视化蛋白质与药物的相互作用。因此，这项工作的结果将为设计具有更好的扩散特性和靶向性的抗生素提供参考。离子型谷氨酸受体离子通道(IGluRs)介导脑和脊髓中绝大多数突触的兴奋性反应。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。这些受体以四聚体的形式组装。然而，以前的晶体结构只显示了二聚体络合物。因此，在第三个项目中，我们解决了一种新型的GluR2 LBD二聚体的晶体结构。