Lipatova T E, Chupryna L M, Pkhakadze G O, Myronov V O
Ukr Biokhim Zh. 1975 Jul-Aug;47(4):532-5.
A method is developed to bind trypsin with the polyurethane matrix. At the first stage the reaction between trypsin and toluylendiisocyanate (TD1) was performed. At the second stage polyisocyanate obtained on the base of TDI-trypsin was sewn with polyoxypropylene glycol with a molecular weight of 1500. 2,4,6-Tris(dimethylaminomethyl)phenol served as an accelerator of the reaction. The polymer obtained is fine-porous and high-elastic. The chemical interaction of trypsin with the polymer is confirmed by the method of IR-spectroscopy and a decrease in the amount of free isocyanate groups. It is shown that the enzymic activity of trypsin chemically bound with the polymer is preserved at a room temperature for several months. When the polymeric net is formed from macrodiisocyanate in the presence of trypsin under conditions when the chemical interaction between the polymer and enzyme activity is also preserved, its value in this case being dependent on the relative content of the enzyme in the polymer -- it is higher in the case when the relative content of the enzyme is less, that may indicate the role of the polymer as a heterogenic cocatalyst or activator. The value of the proteolytic activity of the polymer with the chemically added trypsin is practically constant with its washing off in the column for 24h with 0,05 M veronal buffer whereas under these conditions the polymer with "free" trypsin loses completely the enzymic activity.
开发了一种将胰蛋白酶与聚氨酯基质结合的方法。在第一阶段，进行胰蛋白酶与甲苯二异氰酸酯（TDI）之间的反应。在第二阶段，以TDI-胰蛋白酶为基础得到的多异氰酸酯与分子量为1500的聚氧化丙烯二醇进行缝合。2,4,6-三（二甲氨基甲基）苯酚用作反应促进剂。所得聚合物为细孔且高弹性的。通过红外光谱法和游离异氰酸酯基团数量的减少证实了胰蛋白酶与聚合物之间的化学相互作用。结果表明，与聚合物化学结合的胰蛋白酶的酶活性在室温下可保持数月。当在胰蛋白酶存在下由大分子二异氰酸酯形成聚合物网络时，在聚合物与酶活性之间的化学相互作用也得以保留的条件下，其值在这种情况下取决于聚合物中酶的相对含量——当酶的相对含量较少时该值较高，这可能表明聚合物作为非均相助催化剂或活化剂的作用。用0.05 M巴比妥缓冲液在柱中冲洗24小时后，化学添加胰蛋白酶的聚合物的蛋白水解活性值实际上保持恒定，而在这些条件下，含有“游离”胰蛋白酶的聚合物完全失去酶活性。
