草酸脱羧酶(Oxdc)可用于预防、治疗草酸钙结石症,为了改善其使用性能,该文采用单甲氧基聚乙二醇(mPEG)对Oxdc进行修饰,并对修饰反应条件进行了初步优化,在mPEG-醛/Oxdc摩尔比50∶1,pH 5.0,37℃反应12h条件下,Oxdc修饰率为50.69%,酶活保持率67.52%。经SDS-PAGE和红外光谱(IR)分析,证明mPEG共价结合于酶蛋白分子上。酶学特性分析发现,热处理后修饰草酸脱羧酶(mPEG-Oxdc)的酶活保持21.6%,而野生草酸脱羧酶(Oxdc)酶活完全丧失;在pH 2.0~pH 6.0酸性环境下mPEG-Oxdc的酶活相对高于Oxdc;经胰蛋白酶处理,Oxdc相对酶活仅为20.6%,而mPEG-Oxdc保持47.1%。结果表明:mPEG-Oxdc相对于Oxdc,其热稳定性、pH适应性及对胰蛋白酶的耐受性都有所增强。

In order to improve the operational performance of the oxalate decarboxylase( Oxdc) which can be used to prevent and treat calcium oxalate stone,modification of Oxdc by methoxy polyethyleneglycol was studied, some enzymatic properties of wild enzyme( Oxdc) and modification enzyme( mPEG-Oxdc) had been determined. The modification conditions were preliminary optimized according to the single-factor experiment,and the optimal results were as follows: under a weak acid environment( pH5. 0),a mole ratio of mPEG-aldehyde/Oxdc for 50∶ 1,the modification was occurred at 37℃ for 12 h,which could remain about 67. 52% of enzymatic activity and the modification rate was 50. 69%. SDS-PAGE and FT-IR analysis turned out that the mPEG had covalently bound to the enzymatic protein. Furthermore,analysis of enzymatic properties found that the Oxdc lost almost total of enzyme activity,but the mPEG-Oxdc maintained 21. 6% after heating treatment. The activity of mPEG-Oxdc was always higher than the Oxdc during pH2. 0 ~ pH6. 0. As the activity of Oxdc went down to 20. 6%,the mPEG-Oxdc kept 47. 1% after trypsin treatment. All the results demonstrated that the thermostability,resistance to acid,heat and trypsin of mPEG-Oxdc had been improved relatively.