THEORY OF AQUEOUS TWO-PHASE EXTRACTION

人们对新的蛋白质分离技术的兴趣已经在 近年来，基因工程的最新进展和 医疗部门对高纯度蛋白质的需求增加 和酵素。最需要的是恢复过程，这些过程 对于重组蛋白和构成蛋白来说足够温和 细胞内的蛋白质，但很容易适应大规模 制作。同时满足这两个要求的技术是 含水的聚合物两相体系的萃取 聚乙二醇和葡聚糖，或聚乙二醇和葡聚糖 盐。拟议研究的目的是开发一种 以统计力学为基础的一般、综合理论 蛋白质在含或的两相聚合物水溶液中的溶解度 不加盐。这一理论有望为我们提供对 蛋白质分配的分子机制。这个 目标是预测自由能，水相分离 和蛋白质分配系数作为蛋白质和 聚合物浓度、蛋白质和聚合物分子量， 蛋白质电荷、pH、蛋白质类型、聚合物类型、盐类、离子 亲和配基的强度、温度和结合能。 该理论还应该允许盐分结合的预测和 阳离子分配系数。单蛋白的分配 结果将与关于分区的实验数据进行比较 溶菌酶、富马酸酶、苹果酸脱氢酶、人血清 白蛋白和过氧化氢酶。多重蛋白质分配结果 将根据聚乙烯的实验数据进行测试 含CO-Hb的乙二醇/葡聚糖水溶液和 人血清白蛋白。这项拟议的研究旨在 为组织现有数据提供有用的框架 双水相萃取及其在新产品优化设计中的应用 基于这种方法的生物恢复过程。

Interest in new protein isolation techniques has been stimulated in recent years by recent advances in genetic engineering and by increased demand from the medical sector for high purity proteins and enzymes. Most needed are recovery processes which are gentle enough for recombinant proteins and constitutive intracellular proteins, and yet are easily adapted to large-scale production. A technique which fulfills both requirements is extraction with aqueous polymer two-phase systems containing either polyethylene glycol and dextran, or polyethylene glycol and salt. The aim of the proposed research is the development of a general, comprehensive theory, based in statistical mechanics of protein solubility in two-phase aqueous polymer solutions with or without salt. The theory is expected to provide insight into the molecular mechanisms responsible for protein partitioning. The objective is to predict free energies, aqueous phase separations and protein partition coefficients as a function of protein and polymer concentrations, protein and polymer molecular weights, protein charge, pH, protein type, polymer type, salt type, ionic strength, temperature, and the binding energy of affinity ligands. The theory should also allow the prediction of salt union and cation partition coefficients. The single protein partitioning results will be test against experimental data on the partitioning of lysozyme, fumarase, malate-dehydrogenase, human serum albumin, and catalase. The multiple protein partitioning results will be tested against experimental data on polyethylene glycol/dextran aqueous solutions containing CO-hemoglobin and human serum albumin. The proposed research is intended to provide a useful framework for the organization of existing data aqueous two-phase extraction and for the optimal design of new biorecovery processes based on this method.