Surfactant-Mediated Protein Chromatography

该项目属于分析和表面化学的一般领域，以及分离科学的子领域。这项研究的目标是(A)通过疏水作用层析，在没有变性的情况下溶解和分离蛋白质混合物，以及(B)探索和利用温和表面活性剂引起的层析选择性的变化。含有正辛基葡萄糖苷、CHAPS或Triton X-100的流动相将在低于和高于各自的临界胶束浓度的浓度范围内进行检测。蛋白质保留率与表面活性剂浓度和离子强度的依赖关系将通过对一组蛋白质的等度色层分析来确定。将测定表面活性剂与固定相的结合等温线。表面活性剂对层析效率、分辨率和线性负载能力的影响也将被确定。蛋白质回收率(天然的和全部的)将被确定，柱上变性的迹象将通过测量比活性、圆二色谱或本征荧光来测试。将探索溶剂强度与界面张力测量之间的关系，并将设计出在层析后将表面活性物质与蛋白质分离的简便方法。这些研究的结果将对蛋白质化学和生物技术产生广泛和重大的潜在影响。

This project is in the general area of analytical and surface chemistry and in the subfield of separation science. The objectives of this research are (a) to solubilize and separate, by hydrophobic interaction chromatography, protein mixtures without denaturation, and (b) to explore and exploit changes in chromatographic selectivity due to mild surfactants. Mobile chromatographic phases containing n-octyl glucoside, CHAPS, or Triton X-100 will be examined over a range of concentrations below and above the respective critical micelle concentrations. The dependence of protein retention on surfactant concentration and ionic strength will be determined by isocratic chromatography for an ensemble of proteins. Isotherms for the binding of surfactants to stationary phases will be determined. The effects of surfactants on chromatographic efficiency, resolving power, and linear loading capacity will also be determined. Protein recoveries (native and total) will be ascertained and indications of on-column denaturation will be tested by measurements of specific activity, circular dichroism, or intrinsic fluorescence. The correlation of solvent strength with interfacial tension measurements will be explored and convenient means will be devised for the separation of surfactant from protein following chromatography. The results of these studies will be of broad and significant potential impact in protein chemistry and biotechnology.