Assembly of Cyanobacterial Carboxysomes

蓝藻羧基体的组装

• 批准号: 8522376
• 负责人: Myat T Lin
• 金额: $ 5.22万
• 依托单位: CORNELL UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2013
• 资助国家: 美国
• 起止时间: 2013-07-01 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8522376
• 关键词: Anabolism; Area; Bacteria; Bicarbonate Ion; Bicarbonate Ions; Biochemical Reaction; Biological; Biotechnology; Boxing; C-terminal; Capsid; Carbon Dioxide; Cell Nucleus; Chloroplasts; Complex; Confocal Microscopy; Cyanobacterium; Drug Delivery Systems; Electron Microscopy; Elements; Engineering; Enzymes; Escherichia coli; Eukaryota; Future; Galactosidase; Genes; Genome; Glycine; Goals; Image; Individual; Kinetics; Knowledge; Label; Metabolic; Methods; Molecular; Monitor; Nuclear; Organelles; Organism; Oxygenases; Pathway interactions; Peptide Signal Sequences; Peptides; Plant Leaves; Plants; Plastids; Play; Property; Proteins; Reaction; Regulation; Reporting; Ribulose-Bisphosphate Carboxylase; Role; Serine; Shapes; Structure; Synechococcus; Techniques; Thick; Tissues; Tobacco; Transgenic Organisms; Viral; Work; base; carbonate dehydratase; design; expression vector; macromolecular assembly; metabolic engineering; novel; prevent; promoter; protein expression; public health relevance; research study; sample fixation; vector

DESCRIPTION (provided by applicant): Many bacteria have a sophisticated mechanism for metabolic regulation that involves microcompartments. These bacterial microcompartments are made up of unique protein shells similar in structure to a viral capsid and enclose two or more enzymes in a particular reaction pathway. Carboxysomes from cyanobacteria and chemoauxotrophs are the most well known bacterial microcompartments. Within the last few years, the crystal structures of several shell components have been solved and have led to remarkable progress in the knowledge of subunit assembly and function of individual components. So far, there has been no report of heterologous assemblies of bacterial microcompartments in eukaryotic organisms. This project will target multiple carboxysome proteins into chloroplasts. This study will employ transient expression of foreign proteins fused with fluorescent tags by agroinfiltration technique. Any molecular complex assembled from the expressed proteins will be monitored with confocal microscopy, and the structures formed will be characterized by electron microscopy. In addition, this work will generate stable transgenic lines, in which carboxysome proteins are expressed from either the nucleus or the chloroplast genome. By expressing all necessary carboxysome structural components, this project will attempt to assemble the carboxysome microcompartment shells in eukaryotes. In addition, this study will determine whether it is possible to target foreign proteins to the carboxysome shells. The proposed study is an important step for engineering novel microcompartments and will have potential future applications in areas such as drug delivery, metabolic engineering and biotechnology.

描述(申请人提供)：许多细菌有一种复杂的代谢调节机制，涉及到微室。这些细菌微室由独特的蛋白质壳组成，在结构上类似于病毒衣壳，并在特定的反应途径中包裹两种或更多酶。蓝藻和营养缺陷型细菌的羧基体是最广为人知的细菌微区。在过去的几年里，几种壳层成分的晶体结构已经被解决，并导致对亚单位组装和单个成分的功能的认识取得了显著的进展。到目前为止，还没有关于细菌微室在真核生物中异源组装的报道。该项目将针对叶绿体中的多个羧体蛋白质。本研究将利用农业渗透技术瞬时表达带有荧光标记的外源蛋白。任何由表达的蛋白质组装而成的分子复合体都将用共聚焦显微镜进行监测，形成的结构将用电子显微镜进行表征。此外，这项工作还将产生稳定的转基因品系，在这些品系中，羧体蛋白可以从细胞核或叶绿体基因组中表达。通过表达所有必要的羧体结构成分，该项目将尝试在真核生物中组装羧体微室壳。此外，这项研究将确定是否有可能将外源蛋白靶向羧基体壳。这项拟议的研究是设计新型微隔室的重要一步，将在药物输送、代谢工程和生物技术等领域具有潜在的应用前景。