氨基酰化酶在LDS溶液中的失活与去折叠的比较研究

A Comparison of Inactivation and Unfolding of Aminoacylase during LDS Denaturation

氨基酰化酶在阴离子去圬剂十二烷基硫酸锂（ＬＤＳ）溶液中的失活与去折叠的研究结果表明，在低浓度的ＬＤＳ溶液（０．６ｍｍｏｌ／Ｌ）中变性时，以荧光和紫外差吸收方法监测的酶分子构象尚未发生明显变化。而酶的活力已经大部分或几乎全部丧失。当ＬＤＳ浓度达１．６ｍｍｏｌ／Ｌ时，此时酶分子的构象变化才达到最大程度，在实验使用的ＬＤＳ的浓度范围内，用远紫外ＣＤ光谱监测的二级结构没有发生明显的变化。从上述研究结果，可以认为含锌氨基酰化酶的活性部位也具有相对的柔性。

The unfolding of aminoacylase in lithium dodecylsulfate (LDS)solutions of different concentrations has been studied by following the changes in the ultraviolet absorbance and intrinsic fluoresence. The results show that the denatur ation of the enzyme results in negative peaks at 287nm and 295 nm in the denatured minus native enzyme difference spectrum. The fluorescence emission intensity of the enzyme decreases in LDS solution of incresing concentrations. The inactivition of this enzyme has been followed and compared with the unfolding observed during LDS denaturation. A marked decrease in enzyme activity is already evident at low LDS concentrations (0. 6 mmol)before significant unfolding can be deteceed. In the LDS concentration regions employed in present study,the changes of secondary structure of aminoacylase in the LDS solutions by following the changes of far ultraviolet CD spectra have not been observed. The above results suggest that the active sites of metal enzyme containing Zn￣2+ also are more flexible conformationally than the whole enzyme.