Single-molecule atomic force spectroscopy reveals that DnaD forms scaffolds and enhances duplex melting.
Single-molecule atomic force spectroscopy reveals that DnaD forms scaffolds and enhances duplex melting.
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DOI：
10.1016/j.jmb.2008.01.067
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发表时间：
2008-03-28
影响因子：
5.6
通讯作者：
Soultanas, Panos
中科院分区：
文献类型：
--
作者：
Zhang, Wenke;Machon, Cristina;Orta, Alberto;Phillips, Nicola;Roberts, Clive J.;Allen, Stephanie;Soultanas, Panos
枯草芽孢杆菌DnaD是一种重要的DNA结合蛋白，参与复制和DNA重塑。利用单分子原子力光谱技术研究了DnaD及其结构域与DNA的相互作用。我们的数据表明，DnaD的固定化的双链体DNA的单分子的结合导致显着减少的“端到端”的DNA的距离在浓度依赖性的方式，与以前报道的DnaD诱导的循环支架形成一致。天然DnaD增强DNA链的部分解链。DnaD的C-末端结构域（Cd）与DNA结合并增强部分双链体解链，但不引起DNA成环。Cd介导的解链不如天然DnaD引起的解链有效。N-末端结构域（Nd）对DNA没有显著影响。Nd和Cd的混合物未能重建天然DnaD的DNA成环效应，但产生与Cd自身完全相同的效应，与先前报道的分离结构域形成DNA相互作用支架的失败一致。
The Bacillus subtilis DnaD is an essential DNA-binding protein implicated in replication and DNA remodeling. Using single-molecule atomic force spectroscopy, we have studied the interaction of DnaD and its domains with DNA. Our data reveal that binding of DnaD to immobilized single molecules of duplex DNA causes a marked reduction in the ‘end-to-end’ distance of the DNA in a concentration-dependent manner, consistent with previously reported DnaD-induced looping by scaffold formation. Native DnaD enhances partial melting of the DNA strands. The C-terminal domain (Cd) of DnaD binds to DNA and enhances partial duplex melting but does not cause DNA looping. The Cd-mediated melting is not as efficient as that caused by native DnaD. The N-terminal domain (Nd) does not affect significantly the DNA. A mixture of Nd and Cd fails to recreate the DNA looping effect of native DnaD but produces exactly the same effects as Cd on its own, consistent with the previously reported failure of the separated domains to form DNA-interacting scaffolds.
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