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Transcription-dependent confined diffusion of enzymes within subcellular spaces of the bacterial cytoplasm
BMC Biology ( IF 5.4 ) Pub Date : 2021-09-02 , DOI: 10.1186/s12915-021-01083-4 Daniel A O Rotter 1, 2 , Christoph Heger 1, 2 , Luis M Oviedo-Bocanegra 1, 2 , Peter L Graumann 1, 2
BMC Biology ( IF 5.4 ) Pub Date : 2021-09-02 , DOI: 10.1186/s12915-021-01083-4 Daniel A O Rotter 1, 2 , Christoph Heger 1, 2 , Luis M Oviedo-Bocanegra 1, 2 , Peter L Graumann 1, 2
Affiliation
Knowledge on the localization and mobility of enzymes inside bacterial cells is scarce, but important for understanding spatial regulation of metabolism. The four central enzymes (Rib enzymes) of the riboflavin (RF) biosynthesis pathway in the Gram positive model bacterium Bacillus subtilis have been studied extensively in vitro, especially the heavy RF synthase, a large protein complex with a capsid structure formed by RibH and an encapsulated RibE homotrimer, which mediates substrate-channeling. However, little is known about the behavior and mobility of these enzymes in vivo. We have investigated the localization and diffusion of the Rib enzymes in the cytoplasm of B. subtilis. By characterizing the diffusion of Rib enzymes in live cells using single particle tracking (SPT) we provide evidence for confined diffusion at the cell poles and otherwise Brownian motion. A majority of RibH particles showed clear nucleoid occlusion and a high degree of confined motion, which is largely abolished after treatment with Rifampicin, revealing that confinement is dependent on active transcription. Contrarily, RibE is mostly diffusive within the cell, showing only 14% encapsulation by RibH nanocompartments. By localizing different diffusive populations within single cells, we find that fast diffusion occurs mostly across the nucleoids located in the cell centers, while the slower, confined subdiffusion occurs at the crowded cell poles. Our results provide evidence for locally different motion of active enzymes within the bacterial cytoplasm, setting up metabolic compartmentalization mostly at the poles of cells.
中文翻译:
酶在细菌细胞质的亚细胞空间内的转录依赖性受限扩散
关于细菌细胞内酶的定位和流动性的知识很少,但对于理解新陈代谢的空间调节很重要。对革兰氏阳性模型细菌枯草芽孢杆菌核黄素 (RF) 生物合成途径的四种中心酶 (Rib 酶) 进行了广泛的体外研究,特别是重 RF 合酶,一种由 RibH 和封装的 RibE 同源三聚体,介导底物通道。然而,关于这些酶在体内的行为和流动性知之甚少。我们研究了 Rib 酶在枯草芽孢杆菌细胞质中的定位和扩散。通过使用单粒子跟踪 (SPT) 表征活细胞中 Rib 酶的扩散,我们为细胞极的受限扩散和布朗运动提供了证据。大多数 RibH 颗粒表现出明显的类核闭塞和高度受限的运动,这在用利福平治疗后基本上被消除,表明受限依赖于主动转录。相反,RibE 主要在细胞内扩散,仅显示 14% 的 RibH 纳米隔间封装。通过在单个细胞内定位不同的扩散群体,我们发现快速扩散主要发生在位于细胞中心的类核上,而较慢、受限的亚扩散发生在拥挤的细胞极。
更新日期:2021-09-03
中文翻译:
酶在细菌细胞质的亚细胞空间内的转录依赖性受限扩散
关于细菌细胞内酶的定位和流动性的知识很少,但对于理解新陈代谢的空间调节很重要。对革兰氏阳性模型细菌枯草芽孢杆菌核黄素 (RF) 生物合成途径的四种中心酶 (Rib 酶) 进行了广泛的体外研究,特别是重 RF 合酶,一种由 RibH 和封装的 RibE 同源三聚体,介导底物通道。然而,关于这些酶在体内的行为和流动性知之甚少。我们研究了 Rib 酶在枯草芽孢杆菌细胞质中的定位和扩散。通过使用单粒子跟踪 (SPT) 表征活细胞中 Rib 酶的扩散,我们为细胞极的受限扩散和布朗运动提供了证据。大多数 RibH 颗粒表现出明显的类核闭塞和高度受限的运动,这在用利福平治疗后基本上被消除,表明受限依赖于主动转录。相反,RibE 主要在细胞内扩散,仅显示 14% 的 RibH 纳米隔间封装。通过在单个细胞内定位不同的扩散群体,我们发现快速扩散主要发生在位于细胞中心的类核上,而较慢、受限的亚扩散发生在拥挤的细胞极。