Chemical Engineering Department, School of Engineering, Monash University Malaysia, Jalan Lagoon Selatan, 47500 Bandar Sunway, Selangor, Malaysia.
Chemical Engineering Department, School of Engineering, Monash University Malaysia, Jalan Lagoon Selatan, 47500 Bandar Sunway, Selangor, Malaysia; Arkema Thiochemicals Sdn Bhd, Oasis Ara Damansara, 47301 Petaling Jaya, Selangor, Malaysia.
Int J Biol Macromol. 2023 Dec 31;253(Pt 3):126665. doi: 10.1016/j.ijbiomac.2023.126665. Epub 2023 Sep 7.
Despite extensive studies revealing the potential of cholinium-based ionic liquids (ILs) in protein stabilization, the nature of interaction between ILs' constituents and protein residues is not well understood. In this work, we used a combined computational and experimental approach to investigate the structural stability of a peptide hormone, insulin aspart (IA), in ILs containing a choline cation [Ch] and either dihydrogen phosphate ([Dhp]) or acetate ([Ace]) as anions. Although IA remained stable in both 1 M [Ch][Dhp] and 1 M [Ch][Ace], [Dhp] exhibited a much stronger stabilization effect than [Ace]. Both the hydrophilic ILs intensely hydrated IA and increased the number of water molecules in IA's solvation shell. Undeterred by the increased number of water molecules, the native state of IA's hydrophobic core was maintained in the presence of ILs. Importantly, our results reveal the importance of IL concentration in the medium which was critical to maintain a steady population of ions in the microenvironment of IA and to counteract the denaturing effect of water molecules. Through molecular docking, we confirm that the anions exert the dominant effect on the structure of IA, while [Ch] have the secondary influence. The computational results were validated using spectroscopic analyses (ultra-violet, fluorescence, and circular dichroism) along with dynamic light scattering measurements. The extended stability of IA at 30 °C for 28 days in 1 M [Ch][Dhp] and [Ch][Ace] demonstrated in this study reveals the possibility of stabilizing IA using cholinium-based ILs.
尽管广泛的研究揭示了基于胆碱的离子液体（ILs）在蛋白质稳定化方面的潜力，但 ILs 成分与蛋白质残基之间的相互作用性质仍未得到很好的理解。在这项工作中，我们使用了一种组合的计算和实验方法来研究肽激素胰岛素Aspart（IA）在含有胆碱阳离子[Ch]和二氢磷酸盐([Dhp])或乙酸盐([Ace])作为阴离子的 IL 中的结构稳定性。尽管 IA 在 1 M [Ch][Dhp]和 1 M [Ch][Ace]中都保持稳定，但[Dhp]表现出比[Ace]更强的稳定化作用。两种亲水性 IL 都强烈水合 IA，并增加了 IA 溶剂化壳中的水分子数量。尽管水分子数量增加，但 IA 疏水性核心的天然状态在 IL 存在下得以维持。重要的是，我们的结果揭示了 IL 浓度在介质中的重要性，这对于维持 IA 微环境中离子的稳定群体和抵消水分子的变性效应至关重要。通过分子对接，我们证实阴离子对 IA 的结构具有主导影响，而[Ch]具有次要影响。计算结果通过光谱分析（紫外、荧光和圆二色性）以及动态光散射测量得到验证。本研究中在 30°C 下 28 天内 IA 在 1 M [Ch][Dhp]和[Ch][Ace]中的扩展稳定性表明，使用基于胆碱的 ILs 稳定 IA 是可能的。
