Lipid Nanobilayers to Host Biological Nanopores for DNA Translocations
Lipid Nanobilayers to Host Biological Nanopores for DNA Translocations
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DOI：
10.1021/la3041506
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发表时间：
2013-01-08
期刊：
影响因子：
3.9
通讯作者：
Keyser, Ulrich F.
中科院分区：
文献类型：
--
作者：
Goepfrich, Kerstin;Kulkarni, Chandrashekhar V.;Keyser, Ulrich F.
我们描述了最近引入的一种新的纳米双层技术[Gornall，J.L.；Mahendran，K.R.；Pumbers，O.J.；Steinbock，L.J.；Otto，O.；Chimerel，C.；Winterhalter，M.；Keyser，U.F.在纳米脂双层中蛋白质孔的简单重建。纳米莱特。2011，11(8)，3334-3340]及其纳入金黄色葡萄球菌生物纳米孔α-溶血素的实用方面，以及在各种条件下生物分子移位的后续研究。与传统的双层膜方法相比，该技术具有优势，特别是双层膜的快速形成和扩展的稳定性。我们还开发了一种方法来制备质量均匀的巨型单层囊泡(GUV)，并以可重复的方式生产纳米薄膜。利用各种重要的参数，包括pH、外加电压、盐浓度和纳米孔数，研究了α-溶血素在纳米膜中的重组过程和特征。低pH区α-溶血素残基的质子化影响易位持续时间，这反过来又改变了静电导致的事件类型的统计，并潜在地改变了DNA的结构变化。当改变pH和施加电压时，通过α-溶血素纳米孔研究和部分控制移位事件的捕获速率和类型是可能的。这项研究可能有助于利用纳米薄膜技术进行进一步的探索，特别是因为它与现有的双层方法相比具有优势和技术上的简便性。
We characterize a recently introduced novel nanobilayer technique [Gornall, J. L.; Mahendran, K. R; Pambos, O. J.; Steinbock, L. J.; Otto, O.; Chimerel, C.; Winterhalter, M.; Keyser, U. F. Simple reconstitution of protein pores in nano lipid bilayers. Nano Lett. 2011, 11(8), 3334-3340] and its practical aspects for incorporating the biological nanopore alpha-hemolysin from Staphylococcus aureus and subsequent studies on the translocation of biomolecules under various conditions. This technique provides advantages over classical bilayer methods, especially the quick formation and extended stability of a bilayer. We have also developed a methodology to prepare a uniform quality of giant unilamellar vesicles (GUVs) in a reproducible way for producing nanobilayers. The process and the characteristics of the reconstitution of alpha-hemolysin in nanobilayers were examined by exploiting various important parameters, including pH, applied voltage, salt concentration, and number of nanopores. Protonation of alpha-hemolysin residues in the low pH region affects the translocation durations, which, in turn, changes the statistics of event types as a result of electrostatics and potentially the structural changes in DNA. When the pH and applied voltage were varied, it was possible to investigate and partly control the capture rates and type of translocation events through alpha-hemolysin nanopores. This study could be helpful to use the nanobilayer technique for further explorations, particularly owing to its advantages and technical ease compared to existing bilayer methods.