Characterization of Caulobacter crescentus FtsZ Protein Using Dynamic Light Scattering
Characterization of Caulobacter crescentus FtsZ Protein Using Dynamic Light Scattering
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DOI：
10.1074/jbc.m111.309492
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发表时间：
2012-07-06
影响因子：
4.8
通讯作者：
Garstecki, Piotr
中科院分区：
文献类型：
--
作者：
Hou, Sen;Wieczorek, Stefan A.;Garstecki, Piotr
微管蛋白同源物FtsZ在细胞中间的自组装是细菌细胞分裂的关键步骤。本文引入动态光散射（DLS）光谱作为一种新的方法来研究FtsZ在溶液中的聚合动力学。DLS数据分析表明，FtsZ聚合物在长度上具有明显的单分散性，与GTP、GDP和FtsZ单体的浓度无关。聚合物扩散系数的测量表明，在游离GTP被消耗之前，它们的长度是非常稳定的。我们估计在这个稳定长度区间内，FtsZ聚合物的平均尺寸在9到18个单体之间。FtsZ的聚合和解聚速率可能受到GDP浓度的影响，因为GTP在FtsZ中的反复加入增加了聚合速率，减慢了解聚速率。增加FtsZ浓度对FtsZ聚合物的粒径没有影响；然而，它通过消耗游离GTP来提高解聚反应的速度。利用透射电子显微镜，我们观察到FtsZ在溶液中形成线性聚合物，在与表面接触后迅速转化为大束，时间尺度短至几秒钟。最后，研究中结合FtsZ效果最好的小分子PC190723在体外对新月形根杆菌FtsZ丝状体没有稳定作用，这补充了前人对大肠杆菌FtsZ的研究，证实了这类小分子对革兰氏阴性FtsZ结合较弱。
The self-assembly of the tubulin homologue FtsZ at the mid-cell is a critical step in bacterial cell division. We introduce dynamic light scattering (DLS) spectroscopy as a new method to study the polymerization kinetics of FtsZ in solution. Analysis of the DLS data indicates that the FtsZ polymers are remarkably monodisperse in length, independent of the concentrations of GTP, GDP, and FtsZ monomers. Measurements of the diffusion coefficient of the polymers demonstrate that their length is remarkably stable until the free GTP is consumed. We estimated the mean size of the FtsZ polymers within this interval of stable length to be between 9 and 18 monomers. The rates of FtsZ polymerization and depolymerization are likely influenced by the concentration of GDP, as the repeated addition of GTP to FtsZ increased the rate of polymerization and slowed down depolymerization. Increasing the FtsZ concentration did not change the size of FtsZ polymers; however, it increased the rate of the depolymerization reaction by depleting free GTP. Using transmission electron microscopy we observed that FtsZ forms linear polymers in solutions which rapidly convert to large bundles upon contact with surfaces at time scales as short as several seconds. Finally, the best studied small molecule that binds to FtsZ, PC190723, had no stabilizing effect on Caulobacter crescentus FtsZ filaments in vitro, which complements previous studies with Escherichia coli FtsZ and confirms that this class of small molecules binds Gram-negative FtsZ weakly.