摘要
采用超滤、制备电泳对木瓜蛋白酶进行分离纯化。结果显示:超滤后木瓜蛋白酶的蛋白酶和壳聚糖酶比活力分别从435.10U/mg和2.17U/mg提高到531.39U/mg和2.65U/mg,纯化倍数均提高了1.22倍。制备电泳图谱上显示有五条明显的酶条带,酶活性测定显示条带2具有较高的壳聚糖酶比活力。采用SDS-PAGE和HPLC对此纯化酶组分进行纯度鉴定,结果显示为单一条带和单一峰,说明木瓜蛋白酶中存在具有水解壳聚糖活力的酶组分。HPLC-MS/MS测定该纯化酶组分的氨基酸序列为MRITISGSPG SGTTTLGRSI AEKYSYRYVS AGEVFRGL AK ERNMDLAAFG KIAETDPAID LEIADARQKEI GESSDDIILE GRLAGWMVEN ADLKILLCAS PECRSTRIAA REGLTEKQ AFEMTIERE ACEAGRYMEYYEI DILDFSPYDL ILNSETFSAN ELFAIVDAAVS SLLKRE。
The fraction with chitosanase activity in a commercial papain preparation was prepared by ultra-filtration separation followed by purification by preparation electrophoresis and its amino acid sequence was analyzed. After ultra-filtration separation with a I00000 MWCO membrane, the protease and chitosanase activities of the papain preparation both increased by 1.22 times, from 435.10 to 531.39 U/rag and from 2.17 to 2.65 U/mg, respectively. Five obvious bands were showed on the preparation electrophoresis pattern of retentates and the brand 2 with a molecular weight between 25 and 35 kD had high chitosanase activity and its homogeneity was demonstrated by SDS-PAGE and HPLC, indicating that there was the enzyme fraction with chitosanase activity in the papain preparation. The amino acid sequence of the enzyme fraction was determined by HPLC-MS/MS as follows： MRITISGSPG SGTITLGRSI AEKYSYRYVS AGEVFRGLAK ERNMDLAAFG KIAETDPAID LEIADARQKEI GESSDDIILE GRLAGWMVEN ADLKILLCAS PECRSTRIAA REGLTEKQAF EMTIEREACE AGRYMEYYEI DILDFSPYDL ILNSETFSAN ELFAIVDAAV SSLLKRE.
出处
《食品科学》
EI
CAS
CSCD
北大核心
2009年第17期235-238,共4页
Food Science
基金
国家自然科学基金项目(20576104)
