Sadosky A B, Wilson J W, Steinman H M, Shuman H A
Department of Microbiology, College of Physicians and Surgeons of Columbia University, New York, New York 10032.
J Bacteriol. 1994 Jun;176(12):3790-9. doi: 10.1128/jb.176.12.3790-3799.1994.
Legionella pneumophila, the causative agent of Legionnaires' disease, contains two superoxide dismutases (SODs), a cytoplasmic iron enzyme (FeSOD) and a periplasmic copper-zinc SOD. To study the role of the FeSOD in L. pneumophila, the cloned FeSOD gene (sodB) was inactivated with Tn903dIIlacZ, forming a sodB::lacZ gene fusion. By using this fusion, expression of sodB was shown to be unaffected by a variety of conditions, including several that influence sod expression in Escherichia coli: aeration, oxidants, the redox cycling compound paraquat, manipulation of iron levels in the medium, and the stage of growth. A reproducible twofold decrease in sodB expression was found during growth on agar medium containing charcoal, a potential scavenger of oxyradicals, in comparison with growth on the same medium without charcoal. No induction was seen during growth in human macrophages. Additional copies of sodB+ in trans increased resistance to paraquat. Construction of a sodB mutant was attempted by allelic exchange of the sodB::lacZ fusion with the chromosomal copy of sodB. The mutant could not be isolated, and the allelic exchange was possible only if wild-type sodB was present in trans. These results indicate that the periplasmic copper-zinc SOD cannot replace the FeSOD. The data strongly suggest that sodB is an essential gene and that FeSOD is required for the viability of L. pneumophila. In contrast, Sod- mutants of E. coli and Streptococcus mutans grow aerobically and SOD is not required for viability in these species.
嗜肺军团菌是军团病的病原体，它含有两种超氧化物歧化酶（SOD），一种细胞质铁酶（FeSOD）和一种周质铜锌SOD。为了研究FeSOD在嗜肺军团菌中的作用，用Tn903dIIlacZ使克隆的FeSOD基因（sodB）失活，形成sodB::lacZ基因融合体。通过使用这种融合体，发现sodB的表达不受多种条件的影响，包括几种影响大肠杆菌中sod表达的条件：通气、氧化剂、氧化还原循环化合物百草枯、培养基中铁水平的调控以及生长阶段。与在不含木炭的相同培养基上生长相比，在含有木炭（一种潜在的氧自由基清除剂）的琼脂培养基上生长期间，发现sodB表达可重复性地降低两倍。在人类巨噬细胞中生长期间未观察到诱导现象。反式导入额外的sodB +拷贝可增加对百草枯的抗性。试图通过sodB::lacZ融合体与sodB的染色体拷贝进行等位基因交换来构建sodB突变体。无法分离出突变体，并且只有当反式存在野生型sodB时等位基因交换才有可能。这些结果表明周质铜锌SOD不能替代FeSOD。数据强烈表明sodB是一个必需基因，并且FeSOD是嗜肺军团菌生存所必需的。相比之下，大肠杆菌和变形链球菌的Sod - 突变体在有氧条件下生长，并且在这些物种中生存不需要SOD。
