Identification, subcellular localization and functional interactions of PilMNOWQ and PilA4 involved in transformation competency and pilus biogenesis in the thermophilic bacterium Thermus thermophilus HB27
Identification, subcellular localization and functional interactions of PilMNOWQ and PilA4 involved in transformation competency and pilus biogenesis in the thermophilic bacterium Thermus thermophilus HB27
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嗜热细菌 HB27 中参与转化能力和菌毛生物发生的 PilMNOWQ 和 PilA4 的鉴定、亚细胞定位和功能相互作用
DOI：
10.1111/j.1742-4658.2006.05335.x
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发表时间：
2006
期刊：
影响因子：
--
通讯作者：
B. Averhoff
中科院分区：
文献类型：
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作者：
Judit Rumszauer;Cornelia Schwarzenlander;B. Averhoff
嗜热菌HB27的自然转化系统由至少16个不同的功能蛋白组成，分别由7个不同的基因座编码。在这篇文章中，我们首次对嗜热嗜热菌能力蛋白PilMNOWQ和PilA4进行了生化分析，并证明了这两个基因都是自然转化所必需的。我们鉴定了三种不同形式的PilA4，其中一种表观分子质量为14 kDa，与推导出的蛋白质的表观分子质量相关，18 kDa和23 kDa；最后一种被发现是糖基化的。我们证明PilM、Piln和Pilo位于内膜，而PilW、PilQ和PilA4位于内膜和外膜。这些数据表明，PilMNOWQ和PilA4是跨越内膜和外膜的DNA转位结构的组成部分。我们进一步证明了PilA4和PilQ都与菌毛结构结合。讨论了PilQ和PilA4在DNA易位和菌毛生物发生中的可能功能。比较突变研究表明，PilW或PilQ的突变显著影响另一种蛋白质在外膜上的位置。此外，这些突变体的外膜中不存在PilA4。根据这些发现，我们认为PilW、PilQ和PilA4在外膜组分中稳定定位或积累的能力相互依赖，这与由PilW、PilQ和PilA4组成的外膜DNA转位复合体相一致。
The natural transformation system of the thermophilic bacterium Thermus thermophilus HB27 comprises at least 16 distinct competence proteins encoded by seven distinct loci. In this article, we present for the first time biochemical analyses of the Thermus thermophilus competence proteins PilMNOWQ and PilA4, and demonstrate that the pilMNOWQ genes are each essential for natural transformation. We identified three different forms of PilA4, one with an apparent molecular mass of 14 kDa, which correlates with that of the deduced protein, an 18‐kDa form and a 23‐kDa form; the last was found to be glycosylated. We demonstrate that PilM, PilN and PilO are located in the inner membrane, whereas PilW, PilQ and PilA4 are located in the inner and outer membranes. These data show that PilMNOWQ and PilA4 are components of a DNA translocator structure that spans the inner and outer membranes. We further show that PilA4 and PilQ both copurify with pilus structures. Possible functions of PilQ and PilA4 in DNA translocation and in pilus biogenesis are discussed. Comparative mutant studies revealed that mutations in either pilW or pilQ significantly affect the location of the other protein in the outer membrane. Furthermore, no PilA4 was present in the outer membranes of these mutants. From these findings, we conclude that the abilities of PilW, PilQ and PilA4 to stably localize or accumulate in the outer membrane fraction are strongly dependent on one another, which is in accord with an outer membrane DNA translocator complex comprising PilW, PilQ, and PilA4.