Toward detection of DNA-bound proteins using solid-state nanopores: insights from computer simulations.
Toward detection of DNA-bound proteins using solid-state nanopores: insights from computer simulations.
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DOI：
10.1002/elps.201200164
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发表时间：
2012-12
期刊：
影响因子：
2.9
通讯作者：
Aksimentiev, Aleksei
中科院分区：
文献类型：
--
作者：
Comer, Jeffrey;Ho, Anthony;Aksimentiev, Aleksei
通过全原子分子动力学模拟，我们探索了在薄合成膜中使用纳米孔来检测和鉴定DNA结合蛋白。再现一个典型的实验设置，我们模拟电场驱动的dna结合蛋白通过比蛋白质直径更小的纳米孔的运输。作为模型系统，我们使用特异性和非特异性结合到双链DNA片段的限制性内切酶EcoRI和BamHI，以及通过生物素连接体结合到双链和单链DNA的链亲和蛋白和中性蛋白。我们的模拟阐明了纳米孔诱导蛋白质- dna复合物破裂的分子力学，纳米孔中电泳力对dna -蛋白质键施加的有效力，以及dna -表面相互作用在破裂过程中的作用。我们评估了纳米孔离子电流和由嵌入电极测量的局部静电电位的能力，以报告DNA捕获、DNA结合蛋白捕获和DNA-蛋白键断裂。我们发现，双链DNA上的应变变化可以通过改变纳米孔离子电流和嵌入电极的电位来揭示蛋白质- DNA复合物的破裂。基于我们的模拟结果，我们提出了一种检测DNA结合蛋白的新方法，该方法利用纳米孔中电泳力下的缺口双链剥离。
Through all-atom molecular dynamics simulations, we explore the use of nanopores in thin synthetic membranes for detection and identification of DNA binding proteins. Reproducing the setup of a typical experiment, we simulate electric field-driven transport of DNA-bound proteins through nanopores smaller in diameter than the proteins. As model systems, we use restriction enzymes EcoRI and BamHI specifically and nonspecifically bound to a fragment of double-stranded DNA, and streptavidin and NeutrAvidin proteins bound to double- and single-stranded DNA via a biotin linker. Our simulations elucidate the molecular mechanics of nanopore-induced rupture of a protein–DNA complex, the effective force applied to the DNA-protein bond by the electrophoretic force in a nanopore, and the role of DNA-surface interactions in the rupture process. We evaluate the ability of the nanopore ionic current and the local electrostatic potential measured by an embedded electrode to report capture of DNA, capture of a DNA-bound protein, and rupture of the DNA-protein bond. We find that changes in the strain on double-stranded DNA can reveal the rupture of a protein–DNA complex by altering both the nanopore ionic current and the potential of the embedded electrode. Based on the results of our simulations, we suggest a new method for detection of DNA binding proteins that utilizes peeling of a nicked double strand under the electrophoretic force in a nanopore.