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当前位置: 首页 > 文章 > 发酵乳杆菌细胞壁蛋白酶的分离纯化及酶学性质研究 食品科学 2011,32 (17) 262-268
Position: Home > Articles > Purification and Enzymatic Properties of Cell Envelope Protease from Lactobacillus fermentum FOOD SCIENCE 2011,32 (17) 262-268

发酵乳杆菌细胞壁蛋白酶的分离纯化及酶学性质研究

作  者：
朱姁;潘道东
单  位：
南京师范大学国家乳品加工技术研发分中心;宁波大学生命科学与生物工程学院
关键词：
发酵乳杆菌;细胞壁蛋白酶;纯化;酶学性质
摘  要：
研究分离纯化发酵乳杆菌细胞壁蛋白酶(cell envelope proteinase,CEP)的方法及其酶学性质。用50mmol/LTris-HCl缓冲液(pH7.8)悬浮菌体,进行超声波破碎,细胞质量浓度为0.06g/mL,破碎功率330W,工作220次(工作时间3s,间隔时间5s),离心取上清液即为粗酶液。60%硫酸铵沉淀,Sephacryl S-300 HR凝胶层析,Native-PAGE割胶回收纯化发酵乳杆菌的CEP。用纯化的CEP酶解脱脂乳,酶解液ACE(angiotensin I-converting enzyme)抑制率为50%。SDS-PAGE检测CEP分子质量为32.5kD,最适酶反应温度为41℃,最适酶反应pH值为8.0。Mg2+、Co2+、Ca2+对CEP活性有激活作用;Zn2+、Ni2+、PMSF、EDTA对CEP活性有抑制作用,说明CEP为丝氨酸蛋白酶且酶的活性中心结构的维持与金属离子有关。
译  名：
Purification and Enzymatic Properties of Cell Envelope Protease from Lactobacillus fermentum
作  者：
ZHU Xu 1,PAN Dao-dong1,2, (1.Branch of National Dairy Processing Technology Developing Center,Nanjing Normal University,Nanjing 210097,China; 2.Faculty of Life Science and Biotechnology,Ningbo University,Ningbo 315211,China)
关键词：
Lactobacillus fermentum;cell envelope protease;purification;enzymatic properties
摘  要：
The purification process of cell envelope protease(CEP) from Lactobacillus fermentum was explored in this paper.Cells were suspended in 50 mmol/L Tris-HCl(pH 7.8) and subjected to ultrasonication(cell concentration of 0.06 g/mL,ultrasonic power of 330 W,ultrasonic treatment time of 3 s with intermission of 5 s,and repeated ultrasonic treatment number of 220).The supernatant was collected,precipitated with 60% saturated(NH4)2SO4 solution and separated by Sephacryl S-300 HR chromatography.The active protease was recovered from Native-PAGE gel.The ACE inhibitory rate of purified CEP was 50%.The molecular mass of purified CEP estimated by SDS-PAGE was approximately 32.5 kD.Maximum activity was reached at pH 8.0 and 41℃.The activity of CEP could be activated by Mg2+,Co2+ and Ca2+ and inhibited by Zn2+,Ni2+ and PMSF,suggesting that CEP belongs to serine protease.On the other hand,its activity could also be inhibited by EDTA,suggesting that CEP is a metallopeptidase.

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