Structures of actin-like ParM filaments show architecture of plasmid-segregating spindles.
Structures of actin-like ParM filaments show architecture of plasmid-segregating spindles.
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DOI：
10.1038/nature14356
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发表时间：
2015-07-02
期刊：
影响因子：
64.8
通讯作者：
Loewe, Jan
中科院分区：
文献类型：
--
作者：
Bharat, Tanmay A. M.;Murshudov, Garib N.;Sachse, Carsten;Loewe, Jan
大肠杆菌低拷贝数质粒R1的活性分离涉及由左旋双螺旋肌动蛋白样ParM丝组成的双极纺锤体的形成。ParR将丝与着丝性parC质粒DNA连接起来，同时促进了ParM丝的亚基添加。不断生长的ParMRC纺锤体将姐妹质粒推向细胞极点。在这里，我们使用现代电子低温显微镜方法在体外和细胞中研究了ParM细丝的结构和排列，揭示了亚基和细丝如何在近原子分辨率下结合在一起，产生已知最简单的有丝分裂机制。为了了解动态不稳定性的机制，我们测定了不同核苷酸状态下ParM丝的结构。在4.3 Å分辨率下确定了与AMPPNP结合的细丝的结构并进行了细化。ParM纤维结构表现出较强的纵向界面和较弱的横向相互作用。利用电子低温显微镜，我们重建了形成反平行纺锤体的ParM双峰。最后，通过全细胞电子冷冻断层扫描，我们发现在含有ParMRC位点的低拷贝数质粒的细菌细胞中有大量的重偶体，导致R1质粒分离的异步模型。
Active segregation of E. coli low-copy number plasmid R1 involves formation of a bipolar spindle made of left-handed double-helical actin-like ParM filaments . ParR links the filaments with centromeric parC plasmid DNA, while facilitating the addition of subunits to ParM filaments . Growing ParMRC spindles push sister plasmids to the cell poles . Here, using modern electron cryomicroscopy methods we have investigated the structures and arrangements of ParM filaments in vitro and in cells, revealing at near atomic resolution how subunits and filaments come together to produce the simplest known mitotic machinery. To understand the mechanism of dynamic instability we determined structures of ParM filaments in different nucleotide states. The structure of filaments bound to AMPPNP was determined at 4.3 Å resolution and refined. The ParM filament structure shows strong longitudinal interfaces and weaker lateral interactions. Also using electron cryomicroscopy, we reconstructed ParM doublets forming antiparallel spindles. Finally, with whole-cell electron cryotomography we show that doublets are abundant in bacterial cells containing low-copy number plasmids with the ParMRC locus, leading to an asynchronous model of R1 plasmid segregation.
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影响因子：
48
作者：
Kucukelbir, Alp;Sigworth, Fred J.;Tagare, Hemant D.
通讯作者：
Tagare, Hemant D.
DOI：
10.1107/s0907444904019158
发表时间：
2004-12-01
影响因子：
2.2
作者：
Emsley, P;Cowtan, K
通讯作者：
Cowtan, K
影响因子：
11.4
作者：
Moller-Jensen, Jakob;Ringgaard, Simon;Loewe, Jan
通讯作者：
Loewe, Jan
影响因子：
3
作者：
Kremer, JR;Mastronarde, DN;McIntosh, JR
通讯作者：
McIntosh, JR
影响因子：
3.5
作者：
Izore, Thierry;Duman, Ramona;Loewe, Jan
通讯作者：
Loewe, Jan