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首页> 美国卫生研究院文献>Journal of Bacteriology >Gonococcal MinD Affects Cell Division in Neisseria gonorrhoeae and Escherichia coli and Exhibits a Novel Self-Interaction
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Gonococcal MinD Affects Cell Division in Neisseria gonorrhoeae and Escherichia coli and Exhibits a Novel Self-Interaction

机译：淋球菌MinD影响淋病奈瑟氏球菌和大肠杆菌中的细胞分裂并展现出新型的自我相互作用

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The Min proteins are involved in determining cell division sites in bacteria and have been studied extensively in rod-shaped bacteria. We have recently shown that the gram-negative coccus Neisseria gonorrhoeae contains a min operon, and the present study investigates the role of minD from this operon. A gonococcal minD insertional mutant, CJSD1, was constructed and exhibited both grossly abnormal cell division and morphology as well as altered cell viability. Western blot analysis verified the absence of MinD from N. gonorrhoeae (MinDNg) in this mutant. Hence, MinDNg is required for maintaining proper cell division and growth in N. gonorrhoeae. Immunoblotting of soluble and insoluble gonococcal cell fractions revealed that MinDNg is both cytosolic and associated with the insoluble membrane fraction. The joint overexpression of MinCNg and MinDNg from a shuttle vector resulted in a significant enlargement of gonococcal cells, while cells transformed with plasmids encoding either MinCNg or MinDNg alone did not display noticeable morphological changes. These studies suggest that MinDNg is involved in inhibiting gonococcal cell division, likely in conjunction with MinCNg. The alignment of MinD sequences from various bacteria showed that the proteins are highly conserved and share several regions of identity, including a conserved ATP-binding cassette. The overexpression of MinDNg in wild-type Escherichia coli led to cell filamentation, while overexpression in an E. coli minD mutant restored a wild-type morphology to the majority of cells; therefore, gonococcal MinD is functional across species. Yeast two-hybrid studies and gel-filtration and sedimentation equilibrium analyses of purified His-tagged MinDNg revealed a novel MinDNg self-interaction. We have also shown by yeast two-hybrid analysis that MinD from E. coli interacts with itself and with MinDNg. These results indicate that MinDNg is required for maintaining proper cell division and growth in N. gonorrhoeae and suggests that the self-interaction of MinD may be important for cell division site selection across species.
机译：Min蛋白与确定细菌中的细胞分裂位点有关，并且已经在棒状细菌中进行了广泛的研究。我们最近显示，革兰氏阴性球菌淋病奈瑟氏球菌含有min操纵子，本研究调查了minD在该操纵子中的作用。构建了淋球菌minD插入突变体CJSD1，该突变体显示出严重异常的细胞分裂和形态，以及改变的细胞活力。蛋白质印迹分析证实该突变体中不存在淋病奈瑟氏球菌（MinDNg）的MinD。因此，MinDNg是维持淋病奈瑟菌中适当的细胞分裂和生长所必需的。可溶性和不溶性淋球菌细胞级分的免疫印迹显示，MinDNg既是胞质的，又与不溶性膜级分有关。从穿梭载体联合过量表达MinCNg和MinDNg导致淋球菌细胞显着增大，而仅用编码MinCNg或MinDNg的质粒转化的细胞未显示出明显的形态变化。这些研究表明，MinDNg可能与MinCNg联合抑制淋球菌细胞分裂。来自各种细菌的MinD序列的比对表明，这些蛋白质是高度保守的，并具有多个同一性区域，包括一个保守的ATP结合盒。 MinDNg在野生型大肠杆菌中的过表达导致细胞细丝化，而在大肠杆菌minD突变体中的过表达恢复了大多数细胞的野生型形态。因此，淋球菌MinD在整个物种中都具有功能。酵母两杂交研究以及纯化的带有His标签的MinDNg的凝胶过滤和沉降平衡分析揭示了一种新颖的MinDNg自相互作用。我们还通过酵母双杂交分析表明，来自大肠杆菌的MinD与自身以及与MinDNg相互作用。这些结果表明MinDNg是维持淋病奈瑟菌适当的细胞分裂和生长所必需的，并且表明MinD的自相互作用对于跨物种的细胞分裂位点选择可能很重要。

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