Streamlined Construction of the Cyanobacterial CO2-Fixing Organelle via Protein Domain Fusions for Use in Plant Synthetic Biology
Streamlined Construction of the Cyanobacterial CO2-Fixing Organelle via Protein Domain Fusions for Use in Plant Synthetic Biology
复制标题
DOI：
10.1105/tpc.15.00329
复制
发表时间：
2015-09-01
期刊：
影响因子：
11.6
通讯作者：
Kerfeld, Cheryl A.
中科院分区：
文献类型：
--
作者：
Gonzalez-Esquer, C. Raul;Shubitowski, Tyler B.;Kerfeld, Cheryl A.
细菌微区室（BMC）是一种自组装的细胞器，它将生物化学途径的片段隔离在蛋白质外壳内。鉴于其功能多样性，BMC构成了合成生物学应用的代谢模块的丰富来源。碳氧体，蓝藻BMC的CO2固定，已引起了显着的关注，作为安装到叶绿体的目标，并作为引入其他类型的BMC到植物的基础。羧基体组装涉及至少六种基因产物之间的一系列蛋白质-蛋白质相互作用，以形成代谢核心，外壳围绕其组装。这种复杂性对羧基体或无数功能不同的BMC中的任何一种转移、调节和组装到异源系统中产生了重大挑战。为了克服这一瓶颈，我们构建了一个嵌合蛋白的蓝细菌细长聚球藻，结构和功能取代四个基因产物所需的羧基体形成。该蛋白质是基于羧基体核心中的蛋白质结构域相互作用而设计的。由此产生的流线型的羧基体支持光合作用。这种策略避免了调节多个基因的需要，并降低了异源系统中羧基体组装所需的遗传负荷。更广泛地说，重新工程化的羧基体代表了构建多功能酶核心的结构域融合方法的概念验证，该方法应普遍适用于针对新功能和细胞环境的BMC工程。
Bacterial microcompartments (BMCs) are self-assembling organelles that sequester segments of biochemical pathways within a protein shell. Given their functional diversity, BMCs constitute a rich source of metabolic modules for applications in synthetic biology. The carboxysome, the cyanobacterial BMC for CO2 fixation, has attracted significant attention as a target for installation into chloroplasts and serves as the foundation for introducing other types of BMCs into plants. Carboxysome assembly involves a series of protein-protein interactions among at least six gene products to form a metabolic core, around which the shell assembles. This complexity creates significant challenges for the transfer, regulation, and assembly of carboxysomes, or any of the myriad of functionally distinct BMCs, into heterologous systems. To overcome this bottleneck, we constructed a chimeric protein in the cyanobacterium Synechococcus elongatus that structurally and functionally replaces four gene products required for carboxysome formation. The protein was designed based on protein domain interactions in the carboxysome core. The resulting streamlined carboxysomes support photosynthesis. This strategy obviates the need to regulate multiple genes and decreases the genetic load required for carboxysome assembly in heterologous systems. More broadly, the reengineered carboxysomes represent a proof of concept for a domain fusion approach to building multifunctional enzymatic cores that should be generally applicable to the engineering of BMCs for new functions and cellular contexts.