Structural Basis of Chemoreception

化学感受的结构基础

• 批准号: 8507076
• 负责人: GRANT J JENSEN
• 金额: $ 27.63万
• 依托单位: CALIFORNIA INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-04-01 至 2017-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8507076
• 关键词: 3-Dimensional; Active Sites; Address; Antibiotics; Back; Bacteria; Bacterial Infections; Binding; Biological; Biological Process; Cell model; Cells; Chemoreceptors; Chemotaxis; Chronic; Classification; Collaborations; Diabetes Mellitus; Disease; Electron Microscope; Electrons; Emerging Technologies; Environment; Exhibits; Film; Freezing; Grant; Health; Image; Individual; Industry; Inflammation; Length; Life; Malignant Neoplasms; Maps; Methods; Modeling; Molecular; Monitor; Motor; Nature; Noise; Nutrient; Optics; Organism; Phosphorylation; Phosphotransferases; Play; Positioning Attribute; Process; Proteins; Relative (related person); Resolution; Role; Rotation; Sampling; Signal Transduction; Spectrometry; Structure; Swimming; System; Tomogram; Toxin; Work; X-Ray Crystallography; base; combat; density; detector; dimer; flexibility; human disease; image processing; improved; in vivo; inorganic phosphate; molecular dynamics; mutant; pathogen; prevent; public health relevance; receptor; reconstruction; research study; response

DESCRIPTION (provided by applicant): Bacteria are nearly ubiquitous, play vital roles in industry and the environment, and are important actors in both health and disease for humans and other organisms. They are also small, easily-manipulable model cells that can be used to study basic cell biological studies. Motile bacteria, including many important pathogens, constantly monitor their environment in order to swim towards nutrients and away from toxins, a process called chemotaxis. Attractants and repellents bind to chemoreceptors, which are typically found at the poles of cells grouped together in highly cooperative, ordered arrays. Activated chemoreceptor arrays phosphorylate a protein messenger which in turn binds to flagellar motors, governing the rate of motor reversals and, ultimately, whether the cell continues to move forward or changes direction. While powerful methods like X-ray crystallography and NMR spectrometry have revealed the structures of individual domains of certain chemoreceptors at near-atomic resolution, they have not revealed how the chemoreceptors are arranged inside living cells or the structural basis of array cooperativity. Instead, we have begun to address these issues with an emerging technology, electron cryotomography, which can produce 3-D reconstructions of intact bacterial cells at "macromolecular" (1-5 nm) resolution, which is sufficient to visualize individual receptor dimers. Briefly, bacterial cultures are plunge-frozen in thin films across EM grids and then imaged from a range of angles as the sample is tilted incrementally around one or two axes. 3-D reconstructions are then calculated from the images, and sub-regions with common features can be averaged to increase the signal-to-noise ratio. Following recent work in which we showed that bacterial chemoreceptor arrays are universally arranged in a conserved, 12-nm hexameric lattice of trimers-of-receptor-dimers, here we propose to extend that work in resolution and by imaging fully-activated and -deactivated states. This should reveal how the proteins are arranged within the array as well as the structural basis of activation and array cooperativity. This information will in turn help us understand how bacteria accomplish their roles in health and disease and perhaps suggest new antibiotic targets or strategies.

描述（由申请人提供）：细菌几乎无处不在，在工业和环境中发挥着至关重要的作用，并且是人类和其他生物体健康和疾病的重要参与者。它们也是小型、易于操作的模型细胞，可用于基础细胞生物学研究。运动细菌，包括许多重要的病原体，不断监测其环境，以便游向营养物质并远离毒素，这一过程称为趋化性。引诱剂和驱避剂与化学感受器结合，化学感受器通常位于细胞的两极，这些细胞以高度协作、有序的阵列聚集在一起。激活的化学感受器阵列磷酸化蛋白质信使，该蛋白质信使反过来与鞭毛马达结合，控制运动逆转的速率，并最终控制细胞是否继续前进或改变方向。虽然 X 射线晶体学和核磁共振波谱测定等强大方法已经以近原子分辨率揭示了某些化学感受器的各个域的结构，但它们尚未揭示化学感受器在活细胞内的排列方式或阵列协同性的结构基础。相反，我们已经开始使用新兴技术电子冷冻断层扫描来解决这些问题，该技术可以以“大分子”（1-5 nm）分辨率对完整细菌细胞进行 3D 重建，这足以可视化单个受体二聚体。简而言之，细菌培养物在穿过 EM 网格的薄膜中进行冷冻，然后随着样品围绕一个或两个轴逐渐倾斜，从一系列角度进行成像。然后根据图像计算 3D 重建，并对具有共同特征的子区域进行平均以提高信噪比。最近的研究表明，细菌化学感受器阵列普遍排列在受体三聚体-二聚体的保守的 12 nm 六聚晶格中，在此之后，我们建议通过对完全激活和失活的成像进行分辨率扩展该工作。州。这应该揭示蛋白质在阵列内的排列方式以及激活和阵列协同性的结构基础。这些信息反过来将帮助我们了解细菌如何在健康和疾病中发挥作用，并可能提出新的抗生素靶点或策略。