Sharma S K, Hopkins T R
J Chromatogr. 1975 Jul 16;110(2):321-6. doi: 10.1016/0021-9673(75)85012-6.
The strong and specific binding of chymotrypsin on chromatographic columns containing agarose substituted with N-sigma-amino caproyl-D-tryptophan methyl ester is abolished when the sigma-amino groups on the surface of the enzyme are reacted with acetic anhydride. Because the catalytic properties of the acetylated chymotrypsin are identical to those of the underivatized enzyme, it is concluded that the high affinity of chymotrypsin for this column is not due solely to biospecific inhibitor binding, which is by itself very weak, but requires reinforcement through weak non-specific interactions with the column support. It is postulated that these non-specific interactions include electrostatic interactions between agarose matrix and positively charged lysyl residues on the enzyme. The results demonstrate for the first time that residues on the surface of an enzyme not associated with its active site can play an important role in some chromatographic systems previously thought to be based on purely biospecific interactions.
当用乙酸酐使胰凝乳蛋白酶表面的σ-氨基发生反应时，胰凝乳蛋白酶与含有用N-σ-氨基己酰-D-色氨酸甲酯取代的琼脂糖的色谱柱之间的强而特异性结合就会被消除。由于乙酰化胰凝乳蛋白酶的催化特性与未衍生化的酶相同，因此得出结论，胰凝乳蛋白酶对该色谱柱的高亲和力并非仅归因于本身非常弱的生物特异性抑制剂结合，而是需要通过与色谱柱载体的弱非特异性相互作用来加强。据推测，这些非特异性相互作用包括琼脂糖基质与酶上带正电荷的赖氨酰残基之间的静电相互作用。结果首次证明，酶表面上与其活性位点无关的残基在一些先前认为基于纯生物特异性相互作用的色谱系统中可以发挥重要作用。
