Department of Chemistry, Yale University, New Haven, Connecticut 06520, United States.
Department of Molecular Biophysics and Biochemistry, Yale University, New Haven, Connecticut 06520, United States.
Acc Chem Res. 2023 Jun 20;56(12):1494-1504. doi: 10.1021/acs.accounts.3c00088. Epub 2023 May 10.
Chemists have long been fascinated by chirality, water, and interfaces, making tremendous progress in each research area. However, the chemistry emerging from the interplay of chirality, water, and interfaces has been difficult to study due to technical challenges, creating a barrier to elucidating biological functions at interfaces. Most biopolymers (proteins, DNA, and RNA) fold into macroscopic chiral structures to perform biological functions. Their folding requires water, but water behaves differently at interfaces where the bulk water hydrogen-bonding network terminates. A question arises as to how water molecules rearrange to minimize free energy at interfaces while stabilizing the macroscopic folding of biopolymers to support biological function. This question is central to solving many research challenges, including the molecular origin of biological homochirality, folding and insertion of proteins into cell membranes, and the design of heterogeneous biocatalysts. Researchers can resolve these challenges if they have the theoretical tools to accurately predict molecular behaviors of water and biopolymers at various interfaces. However, developing such tools requires validation by the experimental data. These experimental data are scarce because few physical methods can simultaneously distinguish chiral folding of the biopolymers, separate signals of interfaces from the overwhelming background of bulk solvent, and differentiate water in hydration shells of the polymers from water elsewhere.We recently illustrated these very capacities of chirality-sensitive vibrational sum frequency generation spectroscopy (chiral SFG). While chiral SFG theory dictates that the method is surface-specific under the condition of electronic nonresonance, we show the method can distinguish chiral folding of proteins and DNA and probe water structures in the first hydration shell of proteins at interfaces. Using amide I signals, we observe protein folding into β-sheets without background signals from α-helices and disordered structures at interfaces, thereby demonstrating the effect of 2D crowding on protein folding. Also, chiral SFG signals of C-H stretches are silent from single-stranded DNA, but prominent for canonical antiparallel duplexes as well as noncanonical parallel duplexes at interfaces, allowing for sensing DNA secondary structures and hybridization. In establishing chiral SFG for detecting protein hydration structures, we observe an HO isotopic shift that reveals water contribution to the chiral SFG spectra. Additionally, the phase of the O-H stretching bands flips when the protein chirality is switched from L to D. These experimental results agree with our simulated chiral SFG spectra of water hydrating the β-sheet protein at the vacuum-water interface. The simulations further reveal that over 90% of the total chiral SFG signal comes from water in the first hydration shell. We conclude that the chiral SFG signals originate from achiral water molecules that assemble around the protein into a chiral supramolecular structure with chirality transferred from the protein. As water O-H stretches can reveal hydrogen-bonding interactions, chiral SFG shows promise in probing the structures and dynamics of water-biopolymer interactions at interfaces. Altogether, our work has created an experimental and computational framework for chiral SFG to elucidate biological functions at interfaces, setting the stage for probing the intricate chemical interplay of chirality, water, and interfaces.
化学家长期以来一直对手性、水和界面着迷,在每个研究领域都取得了巨大的进展。然而,由于技术挑战,在手性、水和界面相互作用中出现的化学性质难以研究,这阻碍了人们在界面阐明生物功能。大多数生物聚合物(蛋白质、DNA 和 RNA)折叠成宏观手性结构以发挥生物功能。它们的折叠需要水,但水在界面处的行为却不同,在界面处,体相氢键网络终止。一个问题是,水分子如何在界面处重新排列以最小化自由能,同时稳定生物聚合物的宏观折叠以支持生物功能。这个问题是解决许多研究挑战的核心,包括生物同手性的分子起源、蛋白质折叠和插入细胞膜以及异质生物催化剂的设计。如果研究人员拥有准确预测各种界面下水和生物聚合物分子行为的理论工具,他们就可以解决这些挑战。然而,开发这种工具需要实验数据的验证。由于很少有物理方法可以同时区分生物聚合物的手性折叠,将界面信号与压倒性的体相溶剂背景区分开来,以及区分聚合物水合壳中的水与其他地方的水,因此这些实验数据非常稀缺。我们最近通过手性敏感的振动和频产生光谱(chiral SFG)证明了这些能力。虽然手性 SFG 理论表明,在电子非共振条件下,该方法是表面特异性的,但我们表明,该方法可以区分蛋白质和 DNA 的手性折叠,并在界面处探测蛋白质第一个水合壳中的水结构。我们使用酰胺 I 信号观察到蛋白质折叠成 β-折叠,而没有界面处 α-螺旋和无序结构的背景信号,从而证明了 2D 拥挤对蛋白质折叠的影响。此外,在界面处,C-H 伸缩振动的手性 SFG 信号在单链 DNA 中是无声的,但在典型的反平行双链体以及非典型的平行双链体中是显著的,从而可以检测 DNA 二级结构和杂交。在建立用于检测蛋白质水合结构的手性 SFG 时,我们观察到 HO 同位素位移,这表明水对手性 SFG 光谱有贡献。此外,当蛋白质手性从 L 切换到 D 时,O-H 伸缩带的相位翻转。这些实验结果与我们在真空-水界面上水合 β-折叠蛋白的模拟手性 SFG 光谱一致。模拟进一步表明,总手性 SFG 信号的 90%以上来自蛋白质第一个水合壳中的水。我们得出结论,手性 SFG 信号源于非手性水分子,这些水分子在手性蛋白质周围组装成具有手性的超分子结构,手性是从蛋白质转移过来的。由于水 O-H 伸缩可以揭示氢键相互作用,因此手性 SFG 有望在探测界面上水-生物聚合物相互作用的结构和动态方面发挥作用。总之,我们的工作为手性 SFG 阐明界面上的生物功能建立了实验和计算框架,为探测手性、水和界面之间复杂的化学相互作用奠定了基础。