Requirements for Septal Localization and Chromosome Segregation Activity of the DNA Translocase SftA from Bacillus subtilis
Requirements for Septal Localization and Chromosome Segregation Activity of the DNA Translocase SftA from Bacillus subtilis
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DOI：
10.1159/000450725
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发表时间：
2017-01-01
影响因子：
1.2
通讯作者：
Graumann, Peter L.
中科院分区：
文献类型：
--
作者：
El Najjar, Nina;Kaimer, Christine;Graumann, Peter L.
枯草芽孢杆菌有两种DNA易位酶，它们影响染色体分离的后期阶段：SFTA在隔膜关闭之前分离未分离的DNA，而SpoIIIE拯救隔膜包裹的DNA。我们提供的证据表明，SFTA通过其N-末端47个氨基酸的伸展与分裂机制有关，表明SFTA是通过与分裂机制的一个组成部分的蛋白质-蛋白质相互作用来招募的。在没有前20个氨基酸的情况下，SFTA也被招募到中间细胞，这些氨基酸被认为包含一个膜结合基序。细胞分级实验表明，SFTA在胞质组分中存在，在膜组分中有少量存在，说明SFTA在体内是一种可溶性蛋白质。截短的SFTA结构的表达导致了主导的SFTA缺失表型，即使在截短的蛋白质诱导率非常低的情况下，这表明将非功能单体掺入到SFTA六聚体中会取消功能性。迁移率改变实验和表面等离子体结合研究表明，SFTA以协同方式与DNA结合，当与短核苷酸结合而不是与长片段DNA结合时，ATPase活性较低。(C)2017年S.Karger AG，巴塞尔
Bacillus subtilis possesses 2 DNA translocases that affect late stages of chromosome segregation: SftA separates nonsegregated DNA prior to septum closure, while SpoIIIE rescues septum-entrapped DNA. We provide evidence that SftA is associated with the division machinery via a stretch of 47 amino acids within its N-terminus, suggesting that SftA is recruited by protein-protein interactions with a component of the division machinery. SftA was also recruited to mid-cell in the absence of its first 20 amino acids, which are proposed to contain a membrane -binding motif. Cell fractionation experiments showed that SftA can be found in the cytosolic fraction, and to a minor degree in the membrane fraction, showing that it is a soluble protein in vivo. The expression of truncated SftA constructs led to a dominant sftA deletion phenotype, even at very low induction rates of the truncated proteins, indicating that the incorporation of nonfunctional monomers into SftA hexamers abolishes functionality. Mobility shift experiments and surface plasmon binding studies showed that SftA binds to DNA in a cooperative manner, and demonstrated low ATPase activity when binding to short nucleotides rather than to long stretches of DNA. (C) 2017 S. Karger AG, Basel