Institute of Microbiology, Martin Luther University Halle-Wittenberg, Halle (Saale), Germany.
Institute of Microbiology, Martin Luther University Halle-Wittenberg, Halle (Saale), Germany
J Bacteriol. 2018 Jul 25;200(16). doi: 10.1128/JB.00239-18. Print 2018 Aug 15.
A3(2) is a filamentously growing, spore-forming, obligately aerobic actinobacterium that uses both a copper -type cytochrome oxidase and a cytochrome oxidase to respire oxygen. Using defined knockout mutants, we demonstrated that either of these terminal oxidases was capable of allowing the bacterium to grow and complete its developmental cycle. The genes encoding the complex and the oxidase are clustered at a single locus. Using Western blot analyses, we showed that the oxidase branch is more prevalent in spores than the oxidase. The level of the catalytic subunit, CydA, of the oxidase was low in spore extracts derived from the wild type, but it was upregulated in a mutant lacking the supercomplex. This indicates that cytochrome oxidase can compensate for the lack of the other respiratory branch. Components of both oxidases were abundant in growing mycelium. Growth studies in liquid medium revealed that a mutant lacking the oxidase branch grew approximately half as fast as the wild type, while the oxygen reduction rate of the mutant remained close to that of the wild type, indicating that the oxidase was mainly functioning in controlling electron flux. Developmental defects were observed for a mutant lacking the cytochrome oxidase during growth on buffered rich medium plates with glucose as the energy substrate. Evidence based on using the redox-cycling dye methylene blue suggested that cytochrome oxidase is essential for the bacterium to grow and complete its developmental cycle under oxygen limitation. Respiring with oxygen is an efficient means of conserving energy in biological systems. The spore-forming, filamentous actinobacterium grows only aerobically, synthesizing two enzyme complexes for O reduction, the cytochrome cytochrome oxidase supercomplex and the cytochrome oxidase. We show in this study that the bacterium can survive with either of these respiratory pathways to oxygen. Immunological studies indicate that the oxidase is the main oxidase present in spores, but the oxidase compensates if the oxidase is inactivated. Both oxidases are active in mycelia. Growth conditions were identified, revealing that cytochrome oxidase is essential for aerial hypha formation and sporulation, and this was linked to an important role of the enzyme under oxygen-limiting conditions.
A3(2) 是一种丝状生长、孢子形成、严格需氧的放线菌，它既使用铜型细胞色素 c 氧化酶，也使用细胞色素氧化酶来呼吸氧气。通过使用定义明确的敲除突变体，我们证明这两种末端氧化酶都能够使细菌生长并完成其发育周期。编码 复合体和 氧化酶的基因聚集在单个基因座上。通过 Western blot 分析，我们表明在孢子中， 氧化酶分支比 氧化酶更为普遍。野生型孢子提取物中 氧化酶的催化亚基 CydA 水平较低，但在缺乏 超级复合物的突变体中上调。这表明细胞色素氧化酶可以弥补另一个呼吸分支的缺失。两种氧化酶的成分在生长菌丝中都很丰富。在液体培养基中的生长研究表明，缺乏 氧化酶分支的突变体的生长速度约为野生型的一半，而突变体的氧还原率仍接近野生型，表明 氧化酶主要作用于控制电子流。在缓冲丰富的培养基平板上以葡萄糖作为能源底物生长时，缺乏细胞色素氧化酶的突变体表现出发育缺陷。基于使用氧化还原循环染料亚甲蓝的证据表明，细胞色素氧化酶对于细菌在氧气限制下生长和完成其发育周期是必不可少的。在生物系统中，呼吸氧气是一种有效保存能量的方式。产孢子的丝状放线菌 仅需氧生长，合成两种用于 O 还原的酶复合物，即细胞色素 c 细胞色素氧化酶超级复合物和细胞色素 氧化酶。在这项研究中，我们表明细菌可以通过这两种呼吸途径中的任何一种来适应氧气。免疫研究表明， 氧化酶是孢子中主要的氧化酶，但如果 氧化酶失活， 氧化酶会代偿。两种氧化酶在菌丝体中都有活性。确定了生长条件，揭示了细胞色素氧化酶对于气生菌丝形成和孢子形成是必不可少的，这与该酶在氧气限制条件下的重要作用有关。
