通过相频生成振动光谱揭示聚合物和生物大分子的界面分子级结构
Summary
综合利用,总频率生成(SFG)振动光谱有助于揭示聚合物和生物大分子界面发生的链构体顺序和二次结构变化。
Abstract
作为二阶非线性光学光谱,相和频率生成(SFG)振动光谱已广泛应用于研究各种表面和界面。这种非侵入性光学技术可以提供具有单层或亚单层灵敏度的局部分子级信息。我们在这里提供了如何选择性地检测大分子和生物大分子的埋藏界面的实验方法。并在此基础上讨论了丝纤维素的界面二级结构以及模型短链寡核苷酸双相周围的水结构。前者显示链条重叠或空间约束效应,后者显示保护功能,防止由水的性脊柱上部结构产生的Ca2+离子。
Introduction
总频率生成(SFG)振动光谱的发展可以追溯到沈等人30年前1、2年所做的工作。界面选择性和亚单层灵敏度的独特性使SFG振动光谱学受到物理学、化学、生物学和材料科学等领域大量研究人员的赞赏,3,4 , 5.目前,使用SFG正在研究与表面和界面相关的广泛科学问题,特别是与聚合物和生物大分子相关的复杂界面,如链结构和结构松弛。埋藏聚合物界面,蛋白质二级结构,和界面水结构9,10,11,12,13,14, 15,16,17,18,19,20,21,22,23 24,25,26.
对于聚合物表面和界面,通常通过自旋涂层制备薄膜样品,以获得所需的表面或界面。问题产生是由于信号干扰从两个接口的制备薄膜,这导致不便分析收集的SFG光谱27,28,29 。在大多数情况下,振动信号仅来自单个接口,无论是薄膜/基板还是薄膜/另一种介质,都是可取的。实际上,解决这个问题的方法很简单,即实验性地最大化理想界面的光场,并尽量减少另一个界面的光场。因此,菲涅尔系数或局部场系数需要通过薄膜模型计算,并结合实验结果3、9、10、11进行验证。 12,13,14,15,30。
考虑到上述背景,可以研究一些聚合物和生物界面,以便从分子水平上了解基础科学。在以下,以三个界面问题为例:探测聚聚物(2-羟基乙二醇丙烯酸酯)表面和埋藏界面与基板9,在聚苯乙烯(PS)表面形成丝纤维素(SF)二级结构,水结构周围的模型短链寡核苷酸双工16,21,我们将展示SFG振动光谱如何帮助揭示与基础科学相关的界面分子级结构。
Protocol
1. SFG实验 使用商用皮秒 SFG 系统 (材料表),它基于 Nd:YAG 激光提供脉冲宽度为 ±20 ps 和 50 Hz 频率的基本 1064 nm 光束。 通过使用第二和第三谐波模块,将基本 1064 nm 光束转换为 532 nm 光束和 355 nm 光束。直接引导 532 nm 光束作为输入光束,并通过光学参数生成 (OPG)/光学参数放大 (OPA)生成其他输入中红外 (IR) 光束,覆盖频率范围从 1000 到 4000 厘米-1)差频生成 (…
Representative Results
在协议部分的菲涅尔系数部分,我们已经表明,从理论上讲,一次只选择性地检测一个接口是可行的。在这里,实验,我们确认这种方法基本上是正确的,如图5和图6所示。 图5显示了水侵入后埋藏的界面PHEMA结构,带有+150 nm PHEMA水凝胶膜,图6显示了水中表面结构与±430 nm PHEMA水凝胶膜。面板 A 和 B…
Discussion
为了从分子水平上研究结构信息,SFG 具有其固有的优势(即单层或亚单层灵敏度和界面选择性),可用于研究各种接口,如固体/固体、固体/液体、固体/气体、液体/气体、液体/液体接口。尽管设备维护和光学校准仍然耗时,但回报显著,因为可以在表面和接口上获得详细的分子级信息。
探测聚物(2-羟基乙二甲酸酯)溶液中的表面和埋藏界面:如上文所示,可以调整光场系数。我们?…
Declarações
The authors have nothing to disclose.
Acknowledgements
这项研究得到了国家基础研究重点发展计划(2017YFA0700500)和国家自然科学基金(21574020)的支持。中央高校基础研究基金,由江苏省高等学校重点学术项目开发项目和国家实验生物医学工程示范中心资助教育(东南大学)也受到高度赞赏。
Materials
| 1,2-dipalmitoyl-sn-glycero-3-phosphocholine (DPPC) | Avanti Polar Lipids, Inc. | 850355P-1g | |
| Anhydrous ethanol | Sinopharm Chemical Reagent Co., Ltd | 100092680 | ≥99.7% |
| CaF2 prism | Chengdu YaSi Optoelectronics Co., Ltd. | ||
| Calcium chloride anhydrous | Sinopharm Chemical Reagent Co., Ltd | 10005817 | ≥96.0% |
| deuterated DPPC (d-DPPC) | Avanti Polar Lipids, Inc. | 860345P-100mg | |
| Electromagnetic oven | Zhejiang Supor Co., Ltd | C21-SDHCB37 | |
| Langmuir-Blodgett (LB) trough | KSV NIMA Co., Ltd. | KN 2003 | |
| Lithium bromide anhydrous | Sinopharm Chemical Reagent Co., Ltd | 20056926 | |
| Milli-Q synthesis system | Millipore | Ultrapure water | |
| Plasma cleaner | Chengdu Mingheng Science&Technology Co., Ltd | PDC-MG | Oxygen plasma cleaning |
| Poly(2-hydroxyethyl methacrylate) (PHEMA) | Sigma-Aldrich Co., LLC. | 192066 MSDS | Mw = 300 000 |
| Polystyrene | Sigma-Aldrich Co., LLC. | 330345 MSDS | Mw = 48 kDa and Mn = 47 kDa |
| Silk cocoons | From Bombyx mori | ||
| Single complementary strand of oligonucleotide | Nanjing Genscript Biotechnology Co., Ltd. | H03596 | 5'-CGAAGGCTTCCAGCT-3' |
| Single strand of oligonucleotide | Nanjing Genscript Biotechnology Co., Ltd. | H04936 | 3¢-end modified by cholesterol-triethylene glycol(Chol-TEG) (5¢-GCTTCCGAAGGTCGA-3¢) |
| Sodium carbonate anhydrous | Sinopharm Chemical Reagent Co., Ltd | 10019260 | ≥99.8% |
| Spin-coater | Institute of Microelectronics of the Chinese Academy of Sciences | KW-4A | For the prepartion of ploymer films |
| Step profiler | Veeco | DEKTAK 150 | For the measurement of film thickness |
| Sum frequency generation (SFG) vibrational spectroscopy system | EKSPLA | A commercial picosecond SFG system |
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Citar este artigo
Li, X., Ma, L., Lu, X. Interfacial Molecular-level Structures of Polymers and Biomacromolecules Revealed via Sum Frequency Generation Vibrational Spectroscopy. J. Vis. Exp. (150), e59380, doi:10.3791/59380 (2019).