Interface-resolution domain-domain interactome map of the yeast complexome

酵母复合体的界面分辨率域-域相互作用图

• 批准号: 9918431
• 负责人: Marc Vidal
• 金额: $ 67万
• 依托单位: DANA-FARBER CANCER INST
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-20 至 2023-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9918431
• 关键词: Animal Model; Applications Grants; Architecture; Biochemical; Biological Assay; Biological Process; Biology; Cell model; Cell physiology; Cells; Cellular biology; Communities; Complex; Computer Models; Cryoelectron Microscopy; DNA; Data; Data Set; Generations; Genes; Genotype; Goals; Human; Individual; Laboratories; Literature; Luciferases; Maps; Modeling; Molecular; Molecular Machines; Nature; Open Reading Frames; Organism; Organizational Models; Phenotype; Physiological; Protein Interaction Mapping; Proteins; Proteome; RNA-Protein Interaction; Reaction; Reporter; Resolution; Role; Saccharomyces cerevisiae; Set protein; Structure; Surveys; System; Technology; Testing; Time; Translating; Vision; Work; X-Ray Crystallography; Yeasts; base; improved; innovation; interest; macromolecule; novel; predictive modeling; protein complex; protein function; protein protein interaction; reconstitution; transcriptome

Project Summary/Abstract The physical interactome of an organism, which is the network formed by all physical interactions that can occur in a physiologically relevant dynamic range between all its macromolecules, including protein-protein, DNA- protein, and RNA-protein interactions, is a critical layer to allow us to understand cells from a global point-view. In this grant application, we will focus on a particular aspect of global interactome networks: the formation of all molecular machines or complexes needed to execute all major molecular reactions. While protein complexes are responsible for most biological processes, global models of the organization and architecture of complete sets of protein complexes, or “complexomes”, are still vastly incomplete for most species, including human. We propose to precisely map direct domain-domain interactions between subunits of the yeast complexome with a resolution approaching interface-level resolution. We will systematically fragment all open reading frames corresponding to all ~1,200 genes encoding the ~300 complexes of the yeast complexome and test for direct physical interactions between the corresponding encoded domains using four complementary protein-protein interaction (PPI) assays, one based on the reconstitution of the Gal4 protein in yeast (Gal4-Y2H) and the other three based on the reconstitution of a luciferase protein called NanoLuc in three different expression systems. We will then characterize the sequence requirements of the identified domain-domain interactions and first- generation models of complexes will be generated using interface prediction modeling. The long-term vision of this proposal is to generate binary protein interaction information inside each complex, which, eventually, will be extremely valuable to establish the complete architecture of the yeast complexome. This in turn will allow incorporating dynamic aspects of protein complexes in time and space and generating improving predictive models of genotype-phenotype relationships.

项目概要/摘要 生物体的物理相互作用组，是由所有可能发生的物理相互作用形成的网络 在其所有大分子之间的生理相关动态范围内，包括蛋白质-蛋白质、DNA- 蛋白质以及RNA-蛋白质相互作用是让我们从全局角度理解细胞的关键层。 在本次拨款申请中，我们将重点关注全球相互作用组网络的一个特定方面：所有 执行所有主要分子反应所需的分子机器或复合体。虽然蛋白质复合物 负责大多数生物过程、组织的全局模型和完整的体系结构 对于包括人类在内的大多数物种来说，蛋白质复合物组或“复合体”仍然非常不完整。我们 提出精确绘制酵母复合体亚基之间的直接域-域相互作用 分辨率接近接口级分辨率。我们将系统地分割所有开放阅读框 对应于编码酵母复合体的约 300 个复合物的所有约 1,200 个基因，并测试直接 使用四种互补的蛋白质-蛋白质在相应的编码域之间进行物理相互作用 相互作用 (PPI) 测定，一种基于酵母中 Gal4 蛋白 (Gal4-Y2H) 的重建，另一种基于酵母中 Gal4 蛋白的重建 三种基于在三种不同表达系统中重建称为 NanoLuc 的荧光素酶蛋白。 然后，我们将描述已识别的域-域交互的序列要求，并且首先- 复合体的生成模型将使用界面预测建模来生成。的长远愿景 该提议是在每个复合物内部生成二元蛋白质相互作用信息，最终将被 对于建立酵母复合体的完整结构非常有价值。这反过来将允许 将蛋白质复合物在时间和空间上的动态方面结合起来，并产生改进的预测 基因型-表型关系模型。