Structure and function of proteins in hydrated choline dihydrogen phosphate ionic liquid.
Structure and function of proteins in hydrated choline dihydrogen phosphate ionic liquid.
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DOI：
10.1039/c1cp22965f
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发表时间：
2012-01
期刊：
影响因子：
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通讯作者：
K. D. Weaver;Regina M. Vrikkis;Matthew P Van Vorst;Jonathan Trullinger;R. Vijayaraghavan;D. Foureau;I. McKillop;D. Macfarlane;J. Krueger;G. Elliott
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文献类型：
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作者：
K. D. Weaver;Regina M. Vrikkis;Matthew P Van Vorst;Jonathan Trullinger;R. Vijayaraghavan;D. Foureau;I. McKillop;D. Macfarlane;J. Krueger;G. Elliott
离子液体作为蛋白质和其他生物分子稳定化的有前途的介质正在被深入研究。磷酸二氢胆碱（CDHP）已被确定为这一应用的最有前途的候选人之一。在这项工作中，我们更详细地探讨了CDHP可能对蛋白质的热力学、结构和稳定性产生的影响，包括具有治疗意义的影响。采用微量热法和圆二色性分光光度法（CD）来评估蛋白质溶液在不同浓度的CDHP/水混合物中的热稳定性。观察到溶菌酶和白细胞介素-2的热稳定性与CDHP浓度成比例增加。等温滴定量热法（ITC）用于定量结合相互作用，表明稳定性机制似乎不依赖于CDHP与蛋白质的结合。CD和小角X射线散射（SAXS）分析用于探测由于CDHP的存在而引起的结构变化。小角X射线散射表明，蛋白质表面的电荷效应对蛋白质在离子液体中的稳定性起作用，在25 °C下没有观察到溶菌酶的整体三级构象发生显著变化。然而，在37 °C或更高浓度的CDHP下孵育后，观察到蛋白质结构的微小变化。对蛋白质活性的影响进行了监测，使用浊度测定，和CDHP降低蛋白质活性，但不消除it. Protein溶解度也进行了监测，使用浊度测定，并被发现是成反比的溶液中的CDHP的浓度。
Ionic liquids are being intensely studied as promising media for the stabilization of proteins and other biomolecules. Choline dihydrogen phosphate (CDHP) has been identified as one of the most promising candidates for this application. In this work we have probed in more detail the effects that CDHP may have on the thermodynamics, structure, and stability of proteins, including one of therapeutic interest. Microcalorimetry and circular dichroism spectropolarimetry (CD) were used to assess the thermal stability of protein solutions in CDHP/water mixtures at various concentrations. Increasing thermal stability of lysozyme and interleukin-2 in proportion to CDHP concentration was observed. Isothermal titration calorimetry (ITC) was used to quantify binding interactions, and indicate that the mechanism for stability does not appear to be dependent upon CDHP binding to protein. CD and small angle X-ray scattering (SAXS) analyses were used to probe for structural changes due to the presence of CDHP. SAXS indicates charge effects on the surface of the protein play a role in protein stability in ionic liquids, and no significant alteration of the overall tertiary conformation of lysozyme was observed at 25 °C. However, after incubation at 37 °C or at higher concentrations of CDHP, small changes in protein structure were seen. Effects on protein activity were monitored using turbidity assays, and CDHP decreases protein activity but does not eliminate it. Protein solubility was also monitored using a turbidity assay and was found to be inversely proportional to the concentration of CDHP in solution.