Ghimire Ananta, Kasi Rajeswari M, Kumar Challa V
Department of Chemistry, U-3060, University of Connecticut , Storrs, Connecticut 06269-3060, United States.
J Phys Chem B. 2014 May 15;118(19):5026-33. doi: 10.1021/jp500310w. Epub 2014 May 2.
Rational design of protein-polymer composites and their use, under the influence of the stimulus, for numerous applications requires a clear understanding of protein-polymer interfaces. Here, using poly(acrylic acid) (PAA) and lysozyme as model systems, the binding interactions between these macromolecules were investigated by isothermal titration calorimetry. The binding is proposed to require and be governed by "charge neutralization of the protein/polymer interface" and predicted to depend on solution pH. Calorimetric data show strong exothermic binding of lysozyme to PAA with a molar ΔH and TΔS values of -107 and -95 kcal/mol, respectively, at pH 7 and room temperature. Both ΔH and TΔS decreased linearly with increasing pH from 3 to 8, and these plots had slopes of -17.7 and -17.5 kcal/mol per pH unit, respectively. The net result was that the binding propensity (ΔG) was nearly independent of pH but the binding stoichiometry, surprisingly, increased rapidly with increasing pH from 1 lysozyme binding per PAA molecule at pH 3 to 16 lysozyme molecules binding per PAA molecule at pH 8. A plot of stoichiometry vs pH was linear, and consistent with this result, a plot of ln(average size of the protein/polymer complex) vs pH was also linear. Thus, protonation-deprotonation plays a major role in the binding mechanism. "Charge neutralization" of the lysozyme/PAA interface controls the binding stoichiometry as well as the binding enthalpies/entropies in a predictable fashion, but it did not control the binding affinity (ΔG). The pH dependence of lysozyme binding to PAA, demonstrated here, provides a stimuli-responsive system for protein binding and release from the polymer surface.
在刺激作用下，对蛋白质 - 聚合物复合材料进行合理设计并将其用于众多应用，需要清楚地了解蛋白质 - 聚合物界面。在此，以聚丙烯酸（PAA）和溶菌酶作为模型体系，通过等温滴定量热法研究了这些大分子之间的结合相互作用。据推测，这种结合需要并受“蛋白质/聚合物界面的电荷中和”控制，且预计其取决于溶液的pH值。量热数据表明，在pH值为7和室温条件下，溶菌酶与PAA发生强烈的放热结合，摩尔焓变（ΔH）和摩尔熵变（TΔS）值分别为 -107和 -95千卡/摩尔。随着pH值从3增加到8，ΔH和TΔS均呈线性下降趋势，这些曲线的斜率分别为每pH单位 -17.7和 -17.5千卡/摩尔。最终结果是，结合倾向（ΔG）几乎与pH值无关，但令人惊讶的是，结合化学计量比随着pH值的增加而迅速增加，从pH值为３时每个PAA分子结合1个溶菌酶，增加到pH值为8时每个PAA分子结合16个溶菌酶分子。化学计量比与pH值的关系曲线是线性的，与此结果一致，蛋白质/聚合物复合物平均尺寸的自然对数与pH值的关系曲线也是线性的。因此，质子化 - 去质子化在结合机制中起主要作用。溶菌酶/PAA界面的“电荷中和”以可预测的方式控制结合化学计量比以及结合焓/熵，但它并不控制结合亲和力（ΔG）。本文所展示的溶菌酶与PAA结合的pH依赖性，为蛋白质从聚合物表面的结合和释放提供了一个刺激响应系统。
