Polymer and Biophysics Laboratory, School of Physical Sciences, Jawaharlal Nehru University, New Delhi 110067, India.
J Phys Chem B. 2012 Sep 13;116(36):11065-74. doi: 10.1021/jp3049108. Epub 2012 Aug 30.
This study reports pH dependent stability of protein dispersions of five common proteins, bovine serum albumin (BSA), human serum albumin (HSA), immunoglobulin (IgG), β-lactoglobulin (β-Lg), and gelatin-B (Gel-B), all having isoelectric pH, pI ≈ 5, in room temperature ionic liquid solutions of 1-methyl-3-alkyl (hexyl/octyl) imidazolium chloride (concentration 0-0.2% w/v). Molecular hydrophobicity index, (H-index = hydrophobicity/hydrophilicity) of these molecules spanned the range 0.43-0.87. Electrophoretic characteristics, surface tension data and hydrodynamic size information revealed that IL solutions provide dispersion stability owing to specific protein-IL binding which did not alter their pI values though their surface charge was considerably screened. Change in maximum (ζ(max)) and minimum (ζ(min)) zeta potential values observed at pH ~3 (maximum protonated state) and pH ~8 (maximum deprotonated state) could be described universally as function of IL concentration, c as Δζ(x) = [1 - exp(-ac)] where Δζ(x) is either |(ζ(max) - ζ(w))|/ζ(w) or |(ζ(min) - ζ(w))|/ζ(w), and ζ(w) is the corresponding value in water. Tensiometry data showed two major stages of protein-IL interactions: (i) for c < cmc of IL, the IL molecules selectively bind with imidazolium cation through electrostatic forces forming protein-IL (complex) and (ii) for c> cmc free IL-aggregates begin to form. Similarly, we can define Δγ(x) as either |(γ(max) - γ(w))|/γ(w) at pH 3 or |(γ(min) - γ(w))|/γ(w) at pH 8. Both Δζ(x) and Δγ(x) showed linear dependence with c, Δγ(min, max) (or Δζ(min, max)) = (1 - K(γ) (or K(ζ)) H-index), where the slopes K(ζ) and K(γ) defined intermolecular interactions. Hydrodynamic radii data revealed protein stabilization, circular dichroism spectra implied retention of secondary structures, and Raman spectra confirmed a marginal increase in water structure. Results concluded that selective binding of IL molecules to protein surface in the form of bilayer screen protein surface charge, thereby, contributing to its dispersion stability.
本研究报告了五种常见蛋白质（牛血清白蛋白（BSA）、人血清白蛋白（HSA）、免疫球蛋白（IgG）、β-乳球蛋白（β-Lg）和明胶-B（Gel-B））在室温离子液体 1-甲基-3-烷基（己基/辛基）氯化咪唑中的 pH 依赖性稳定性，这些蛋白质的等电点（pI）均约为 5。这些分子的分子疏水性指数（H-index = 疏水性/亲水性）范围为 0.43-0.87。电泳特性、表面张力数据和流体力学尺寸信息表明，IL 溶液通过特定的蛋白质-IL 结合提供了分散稳定性，尽管其表面电荷被大大屏蔽，但并未改变它们的 pI 值。在 pH3（最大质子化状态）和 pH8（最大去质子化状态）下观察到的最大（ζ（max））和最小（ζ（min））zeta 电位值的变化可以普遍描述为 IL 浓度 c 的函数，Δζ（x）= [1 - exp（-ac）]，其中 Δζ（x）是 |（ζ（max）-ζ（w））|/ζ（w）或 |（ζ（min）-ζ（w））|/ζ（w），ζ（w）是相应的水值。张力计数据显示了蛋白质与 IL 相互作用的两个主要阶段：（i）对于 c<IL 的 cmc，IL 分子通过静电相互作用选择性地与咪唑阳离子结合形成蛋白质-IL（复合物），（ii）对于 c>cmc，游离 IL 聚集物开始形成。同样，我们可以将 Δγ（x）定义为 pH 3 时的 |（γ（max）-γ（w））|/γ（w）或 pH 8 时的 |（γ（min）-γ（w））|/γ（w）。Δζ（x）和 Δγ（x）均与 c 呈线性关系，Δγ（min，max）（或 Δζ（min，max））=（1-K（γ）（或 K（ζ））H-index），其中斜率 K（ζ）和 K（γ）定义了分子间相互作用。流体力学半径数据表明蛋白质得到稳定，圆二色光谱表明二级结构得以保留，拉曼光谱证实水结构略有增加。结果表明，IL 分子以双层形式选择性结合到蛋白质表面，从而屏蔽蛋白质表面电荷，从而有助于其分散稳定性。
