Molecular imprinting of proteins and other macromolecules resulting in new adsorbents
Molecular imprinting of proteins and other macromolecules resulting in new adsorbents
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蛋白质和其他大分子的分子印迹产生新的吸附剂
DOI：
10.1002/bit.260390209
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发表时间：
1992
影响因子：
3.8
通讯作者：
A. Klibanov
中科院分区：
文献类型：
--
作者：
K. Dabulis;A. Klibanov
当将模型蛋白牛血清白蛋白（BSA）溶解在多功能配体L-苹果酸的浓缩水溶液中时，将溶液冻干，并用四氢呋喃彻底洗涤固体残余物以提取苹果酸，然后所得（“印迹”）蛋白能够结合26.4 ±0.9摩尔当量的无水乙酸乙酯中的配体。非印迹BSA（即，除了不存在苹果酸之外，以相同方式制备的）在相同条件下结合不到该量的十分之一。此外，印迹和非印迹BSA在水中几乎没有表现出L-苹果酸的结合。即使在真空下长时间孵育后，印迹BSA仍保留其对乙酸乙酯中配体的“记忆”;然而，在水中溶解消除了印迹蛋白的结合能力。L-苹果酸印迹的BSA不仅在乙酸乙酯中而且在许多其他无水溶剂中显示出对该配体的亲和性。发现溶剂形成氢键的倾向越高，其中的蛋白质-配体结合越低，因此指出氢键是这种结合的驱动力。对完全或部分裂解的BSA、其他球状蛋白、谷胱甘肽和聚（L-天冬氨酸）的研究表明，压印性的关键要求是存在足够长的聚合物链。此外，除了蛋白质之外，许多形成氢键的大分子，如葡聚糖及其衍生物，部分水解淀粉和聚（甲基丙烯酸），也可以在乙酸乙酯中进行印迹以随后结合。从这些实验中推断的印迹和结合的机制涉及每个配体分子在水中与聚合物链上的两个或更多个位点的多点氢键合，从而将后者的一段折叠成配体周围的空腔;在冻干和提取配体之后，空腔保留在有机溶剂中（但不在水中）并引起配体结合。这一结论得到了许多苹果酸类似物和相关配体与L-苹果酸印迹BSA结合的结果的支持。最后，牛血清白蛋白与苹果酸印迹作为一个选择性吸附剂的色谱分离的等摩尔混合物的马来酸和丙烯酸在乙酸乙酯。
When the model protein bovine serum albumin (BSA) was dissolved in a concentrated aqueous solution of the multifunctional ligand L‐malic acid, the solution was lyophilized, and the solid residue thoroughly washed with tetrahydrofuran to extract malic acid, then the resultant (“imprinted”) protein was capable of binding 26.4 ±0.9 mol equivalents of the ligand in anhydrous ethyl acetate. The nonimprinted BSA (i.e., that prepared in the same manner apart from the absence of malic acid) bound less then one‐tenth of that amount under identical conditions. Furthermore, both imprinted and nonimprinted BSA exhibited little binding of L‐malic acid in water. The imprinted BSA retained its “memory” for the ligand in ethyl acetate even after a prolonged incubation under vacuum; dissolution in water, however, eliminated the imprinted protein's binding capacity. The BSA imprinted with L‐malic acid displayed affinity for this ligand not only in ethyl acetate but also in many other anhydrous solvents. It was found that the higher the solvent's propensity to form hydrogen bonds, the lower the protein–ligand binding in it, thus pointing to hydrogen bonds as the driving force of this binding. Studies with completely or partially cleaved BSA, with other globular proteins, glutathione, and poly(L‐aspartic acid) revealed that the critical requirement for the imprintability is the presence of a sufficiently long polymeric chain. Moreover, many hydrogen‐bond‐forming macromolecules other than proteins, such as dextrans and their derivatives, partially hydrolyzed starch, and poly(methacrylic acid), also could be imprinted for subsequent binding in ethyl acetate. The mechanism of imprinting and binding inferred from these experiments involves a multipoint hydrogen bonding in water of each ligand molecule with two or more sites on the polymeric chain, thereby folding a segment of the latter into a cavity around the ligand; following lyophilization and extraction of the ligand, the cavities remain in organic solvents (but not in water) and give rise to ligand binding. This conclusion is supported by the results of binding of numerous malic acid analogs and related ligands to BSA imprinted with L‐malic acid. Finally, BSA imprinted with malic acid was used as a selective adsorbent for a chromatographic separation of an equimolar mixture of maleic and acrylic acids in ethyl acetate.