凝胶层析和离子交换层析结合法纯化高盐发酵液中谷氨酰胺转胺酶

Purification of Transglutaminase from High Salt Culture by Gel and Cation Exchange Chromatography

采用凝胶层析和离子交换层析相结合的方法分离纯化高盐培养基中的谷氨酰胺转胺酶,优化的凝胶层析的条件,上样量6 mL,流速为0.25 mL/min;离子交换层析的上样量50 mL,流速为3 mL/min。酶被纯化了4.22倍,比活力达17.33 U/mg蛋白,回收率为77.5%。液相色谱-串联质谱鉴定、蛋白质数据库比对结果表明,纯酶与AAN01353是同种蛋白质。

To purify the transglutaminase from high-salt medium,gel filtration combined with ion exchange chromatography method was employed.Two-step chromatographic conditions（flow rate and the sample volume） were optimized.The final sample volume and a flow rate for Gel chromatography were 6 mL and 0.25 mL/min,respectively.Conditions for ion-exchange chromatography were a sample volume of 50 mL and flow rate of 3 mL/min.Enzyme was purified 4.22-fold,the specific activity and the recovery rate were 17.33 U/mg protein and 77.5%,respectrively.A comparison between liquid chromatography/tandem mass spectrometric identification and the protein database was carried out and the results showed that the purified enzyme had high homology with AAN01353.