Single particle reconstruction

单粒子重建

• 批准号: 7244745
• 负责人: FREDERICK J SIGWORTH
• 金额: $ 27.4万
• 依托单位: HARVARD MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-04-01 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7244745
• 关键词: Affect; Algorithms; Cell physiology; Cells; Condition; Data; Data Set; Development; Electron Microscope; Ensure; Image; Individual; Ion Channel; Laboratories; Macromolecular Complexes; Maps; Methods; Modeling; Molecular Machines; Molecular Structure; Noise; Pharmaceutical Preparations; Potassium Channel; Probability; Property; Proteins; Resistance; Resolution; Shapes; Signal Transduction; Simulate; Statistical Models; Structure; Techniques; Testing; Therapeutic Intervention; Transferrin; Transferrin Receptor; base; density; expectation; macromolecule; particle; programs; reconstruction; theories; three-dimensional modeling

Cryo-EM is a method for determing the structure of the macromolecules and molecular machines that are responsible for cellular function. Cryo-EM single-particle reconstruction (SPR) is the difficult task of deducing the three-dimensional "density" map of a macromolecular "particle" from a set of very noisy electron-microscope images. SPR fails in the cases that the particles are too small, when the images are too noisy, or when high resolution is sought. It fails because under these conditions the orientation of the particles giving rise to the individual images cannot be determined uniquely. We will develop an SPR algorithm that makes use of Robust Estimation theory to more reliably perform reconstructions from challenging datasets. Our algorithm will maximize a clearly-defined statistical quantity (related to the a posteriori probability) while being resistant to the effects of "outlier" images. We will test the algorithm on simulated datasets and on two experimental datasets from small particles. One of these will be the transferrin-transferrin receptor dataset from the Walz lab which provided a successful SPR by conventional techniques; the other will be data acquired from solubilized Slack (hslo2.1) ion channels, for which conventional SPR has to date not been successful. Discerning the structure (detailed shape) of the molecular machines of cells is essential to the understanding of their function, and to the development of therapeutic interventions including drugs that affect their function. Singe-particle cryo-EM is a powerful and flexible method for the determination of structures, but in many cases it fails to provide results, or the results it provides are incorrect. We seek to increase the reliability and usefulness of this method.

Cryo-EM是一种确定大分子和分子机器结构的方法， 负责细胞功能。Cryo-EM单粒子重建（SPR）是一项艰巨的任务， 从一组非常嘈杂的图像中推导出大分子“粒子”的三维“密度”图， 电子显微镜图像。SPR在颗粒太小的情况下失败，当图像 噪声太大，或者当寻求高分辨率时。它失败了，因为在这些条件下， 不能唯一地确定产生各个图像的粒子。我们将开发一个SPR 算法，利用鲁棒估计理论更可靠地执行重建， 挑战数据集我们的算法将最大化一个明确定义的统计量（与a相关 后验概率），同时抵抗“离群”图像的影响。我们将测试算法， 模拟数据集和两个小颗粒的实验数据集。其中之一将是 来自Walz实验室的转铁蛋白-转铁蛋白受体数据集，其通过常规SPR提供了成功的SPR。 技术;另一个将是从溶解的Slack（hslo2.1）离子通道获得的数据，其中 传统的SPR至今没有成功。 辨别细胞分子机器的结构（详细形状）对于理解 这些研究有助于了解它们的功能，并有助于开发包括影响其功能的药物在内的治疗干预措施。 单粒子冷冻电镜是一种强大而灵活的结构测定方法，但在许多情况下， 它不能提供结果，或者它提供的结果不正确。我们力求提高可靠性， 这种方法的实用性。