利用布朗動態法研究電場與離子強度對DNA吸附於脂雙層上電泳行為之影響

Abstract

脂雙層因其與生物分子的高相容性，被廣泛應用於研究中。許多學者利用脂雙層作為平台，來操作和觀察DNA等生物分子的行為。在脂雙層上進行電泳，有助於研究生物分子的動態特性及其在不同環境下的反應，這對於了解細胞膜的物理化學特性、生物分子間的相互作用，以及開發新型醫療檢測技術具有重要意義。先前的實驗結果顯示，當DNA在脂雙層上電泳時，電泳遷移率會隨著直流電場和溶液離子強度的增加而上升，這與DNA在自由溶液中電泳時完全不同。因此，我們推論這些現象和DNA與脂雙層之間的相互作用有關，但實驗無法清楚揭示這些現象的機制，數值模擬成為理解這些機制的最佳方法。 本實驗室先前利用布朗動態法(Brownian dynamics, BD)模擬並分析DNA於脂雙層上的擴散行為，不但重現實驗中觀察的現象，並證實次擴散現象是由基材表面不平整引起的位能井造成的。接下來，我們將利用此模擬系統探討DNA在脂雙層上電泳的非預期現象。為了理解DNA與脂雙層之間的相互作用，我們先模擬了DNA在脂雙層上的電泳，發現電泳遷移率也會隨著直流電場和溶液離子強度增加而上升，與實驗結果一致。在成功重現這些實驗結果後，我們觀察DNA與脂雙層的互動情形。模擬結果顯示，當電場強度上升時，脂雙層中的正電脂質逐漸出現分佈不均勻的現象，且DNA部分鏈段在正電脂質密度較低的地方出現脫附。 因此，我們利用網格分析法計算正電脂質密度的變異數，量化其離散程度。結果顯示，隨著直流電場與溶液離子強度增加，正電脂質的密度變異數上升，且DNA鏈段脫離脂雙層靜電位能範圍的平均程度增加。為了更聚焦在對DNA產生靜電位能的帶正電脂質，我們計算了吸附於DNA上的正電脂質數目，發現該數目會隨著直流電場與溶液離子強度上升而下降。另外，由於帶負電的DNA與正電脂質移動方向相反，當電場強度和溶液離子強度上升時，吸附於DNA上的正電脂質數目減少，我們認為這會導致DNA受到正電脂質反向的拖曳力下降，進而使DNA電泳速度上升。因此，我們計算了吸附於DNA上的正電脂質數目與DNA電泳速度隨時間的變化，發現它們具有高度的負相關性。這代表電泳遷移率為電場的函數，是因為吸附於DNA上的正電脂質數目隨電場改變所造成的。 最後，為理解正電脂質分布不均的原因，我們計算了正電脂質的佩克萊數(Péclet number,Pe)。結果顯示，隨著直流電場強度增加，正電脂質佩克萊數上升。當電場強度使佩克萊數遠大於1時，流動的效果超過擴散的效應，導致正電脂質在脂雙層中分佈不均，模擬結果顯示佩克萊數上升，正電脂質的密度變異數就會越高。

Lipid bilayers are widely used as platforms to observe and manipulate the behavior of biomolecules such as DNA and proteins. Conducting electrophoresis of biomolecules on lipid bilayers helps studying the dynamic properties of biomolecules and their responses to electric field. This is crucial for understanding cell membrane properties, biomolecular interactions, and developing new applications. Previous experimental results showed that DNA electrophoretic mobility on lipid bilayers increases with the DC electric field and solution ionic strength. These phenomena are very different from those in free solutions and not yet understood. We speculated that these phenomena are related to the interactions between DNA and the lipid bilayers. However, the mechanisms could not be elucidated through experiments alone, making molecular simulations an optimal method for understanding these mechanisms. Our laboratory has previously used Brownian dynamics (BD) simulations to investigate the anaomalous DNA diffusion on lipid bilayers. In this research, we further utilized the same simulation system to investigate unexpected phenomena in DNA electrophoresis on lipid bilayers. We first simulated DNA electrophoresis on the bilayer and found that simulated electrophoretic mobility increases with the DC electric field and solution ionic strength, consistent with experimental results. Along with the increasing DNA mobility, we also observed that the positively charged lipids in the bilayer exhibited increasingly uneven distribution, and some segments of the DNA detached from regions with low positive lipid density. We quantified the inhomogeneity of positively charged lipid distribution and found it increases with the electric field and ionic strength, leading to greater degree of DNA detachment. As DNA segments detach from lipid bilayers, the number of positively charged lipids adsorbed onto DNA decreases. The reduction in adsorbed lipids reduces the drag force applied on DNA, causing the increase of DNA electrophoretic mobility. Finally, to understand the cause of the uneven distribution of positively charged lipids, we calculated the Péclet number (Pe) for the positive lipids. The results indicated that with an increase in DC electric field strength, the Péclet number rose. When the electric field strength increased such that the Péclet number was significantly greater than 1, the convective effects dominated over diffusion, leading to an uneven distribution of positively charged lipids in the lipid bilayer. The simulation results showed that as the Péclet number increased, so did the variance in positive lipid density.