Cytochrome bd Oxidase Has an Important Role in Sustaining Growth and Development of Streptomyces coelicolor A3(2) under Oxygen-Limiting Conditions
Cytochrome bd Oxidase Has an Important Role in Sustaining Growth and Development of Streptomyces coelicolor A3(2) under Oxygen-Limiting Conditions
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DOI：
10.1128/jb.00239-18
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发表时间：
2018-08-01
影响因子：
3.2
通讯作者：
Sawers, R. Gary
中科院分区：
文献类型：
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作者：
Fischer, Marco;Falke, Doerte;Sawers, R. Gary
天蓝色链霉菌A3(2)是一种丝状生长、芽胞形成、需氧放线杆菌，它同时使用铜AA(3)型细胞色素C氧化酶和细胞色素BD氧化酶来呼吸氧气。使用定义的基因敲除突变体，我们证明了这两种末端氧化酶中的任何一种都能够允许细菌生长并完成其发育周期。编码BCC复合体和AA(3)氧化酶的基因聚集在一个基因座上。通过Western印迹分析，我们发现BCC-AA(3)氧化酶分支在孢子中比BD氧化酶更普遍。BD氧化酶的催化亚基CydA在野生型孢子提取物中水平较低，但在缺乏BCC-AA(3)超复合体的突变体中表达上调。这表明细胞色素BD氧化酶可以弥补另一呼吸分支的缺失。生长菌丝体中含有丰富的两种氧化物酶成分。液体培养中的生长研究表明，缺失BCC-AA(3)氧化酶分支的突变体的生长速度约为野生型的一半，而氧还原速率与野生型接近，表明BD氧化酶主要控制电子通量。一株缺乏细胞色素BD氧化酶的突变体在以葡萄糖为能量底物的富营养缓冲平板上生长过程中观察到发育缺陷。使用氧化还原循环染料亚甲基蓝的证据表明，细胞色素BD氧化酶是细菌在氧气限制下生长和完成发育周期所必需的。重要的是氧气呼吸是生物系统中保存能量的有效手段。产孢子的丝状天蓝色放线杆菌只能在需氧条件下生长，合成两种用于氧还原的酶复合体，即细胞色素BCC-AA(3)细胞色素氧化酶超复合体和细胞色素BD氧化酶。我们在这项研究中表明，细菌可以通过这两条呼吸途径中的任何一条来呼吸氧气。免疫学研究表明，BCC-AA(3)氧化酶是孢子中存在的主要酶，但如果BCC-AA(3)酶失活，BD氧化酶就会补偿。这两种氧化酶在菌丝体中都很活跃。对生长条件进行了鉴定，揭示了细胞色素BD氧化酶是气生菌丝形成和产孢子所必需的，这与该酶在限氧条件下的重要作用有关。
Streptomyces coelicolor A3(2) is a filamentously growing, spore-forming, obligately aerobic actinobacterium that uses both a copper aa(3)-type cytochrome c oxidase and a cytochrome bd 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 bcc complex and the aa(3) oxidase are clustered at a single locus. Using Western blot analyses, we showed that the bcc-aa(3) oxidase branch is more prevalent in spores than the bd oxidase. The level of the catalytic subunit, CydA, of the bd oxidase was low in spore extracts derived from the wild type, but it was upregulated in a mutant lacking the bcc-aa(3) supercomplex. This indicates that cytochrome bd 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 bcc-aa(3) 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 bd oxidase was mainly functioning in controlling electron flux. Developmental defects were observed for a mutant lacking the cytochrome bd 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 bd oxidase is essential for the bacterium to grow and complete its developmental cycle under oxygen limitation.IMPORTANCE Respiring with oxygen is an efficient means of conserving energy in biological systems. The spore-forming, filamentous actinobacterium Streptomyces coelicolor grows only aerobically, synthesizing two enzyme complexes for O-2 reduction, the cytochrome bcc-aa(3) cytochrome oxidase supercomplex and the cytochrome bd oxidase. We show in this study that the bacterium can survive with either of these respiratory pathways to oxygen. Immunological studies indicate that the bcc-aa(3) oxidase is the main oxidase present in spores, but the bd oxidase compensates if the bcc-aa(3) oxidase is inactivated. Both oxidases are active in mycelia. Growth conditions were identified, revealing that cytochrome bd oxidase is essential for aerial hypha formation and sporulation, and this was linked to an important role of the enzyme under oxygen-limiting conditions.