天冬氨酸酶的分离提纯及其变性复性的研究

以E.Coli J-3为原料,经菌体破碎、链霉素处理、热处理、硫铵分级、磷酸钙胶处理、DEAESephadex A-50柱层析、羟基磷灰石柱层析、Ultrogel AcA 34凝胶过滤、HPLC层析九个步骤,得到聚丙烯酰胺梯度凝胶电泳纯的天冬氨酸酶。并探讨了各种环境因素对变性复性程度的影响。结果表明,不同的变性、复性条件会诱导不同构象的产生。诱导酶的性质与天然酶不同。

The Purification of Aspartase and Its Denaturation and Renaturation

Aspartase was highly purified from Escherichia coli J-3 cells. The purification procedures consisted of sonic extraction, streptomycin treatment, (NH_4)_2SO_4 faction, heat treatment, calcium phosphate gel treatment, column chromatography on DEAE-sephadex, hydroxylapatite, Ultragel AcA34 and high performance liquid chromatography. The purified enzyme preparations were homogeneous as judged by polyacrylamide gradient gel electrophoresis. The influence of a variety of factors on the denaturation and renaturation of aspartase was studied. It has been found that different conditions of denaturation and renaturation can induce different conformations of aspartase. The properties of the renatured enzyme are different from those of native enzyme.