嗜热子囊菌光孢变种Thermoascus aurantiacus var.levisporus内切β-葡聚糖酶的分离纯化及特性  

Purification and Characterization of Endoglucanase from Thermoascus aurantiacus var.levisporus

作　　者：王冬梅[1] 孟军 李多川[3] 

机构地区：[1]莱芜职业技术学院,山东莱芜271100 [2]莱芜市畜牧局,山东莱芜271100 [3]山东农业大学环境生物系,山东泰安271018

出　　处：《中国生物化学与分子生物学报》2009年第10期926-931,共6页Chinese Journal of Biochemistry and Molecular Biology

基　　金：国家高技术研究发展计划资助项目(863计划;No.2003AA241161);国家自然科学基金资助项目(No.3017001330270013)~~

摘　　要：采用培养分离、室内测定等方法,对嗜热子囊菌光孢变种Thermoascus aurantiacus var.levisporus产生的内切β-葡聚糖酶进行了分离纯化及特性研究.粗酶液经硫酸铵沉淀、DEAE-Sepharose Fast Flow阴离子层析、Phenyl-Sepharose疏水层析等步骤获得了凝胶电泳均一的内切β-葡聚糖酶.结果表明,经12%SDS-PAGE测得酶的单亚基分子量约为31.5 kD,凝胶过滤层析测得酶的分子量约为34.5 kD.该酶反应的最适温度为55℃,最适pH为2.5～3.0.该酶在pH3.0条件下60℃较为稳定;80℃保温30 min有20%原酶活性.金属离子对内切β-葡聚糖酶活性影响较大,其中K+、Ca2+、Mn2+对酶有激活作用;Ag+、Cu2+、Al3+对酶有显著抑制作用.该酶对羧甲基纤维素具有很强的底物特异性.Endoglucanase was purified from culture supernatant of Thermoascus aurantiacus var. levisporus to homogeneity, using ammonium sulfate fraetionation, DEAE-Sepharose, Phenyl-Sepharos, and Sephacryl S- 100 fast-flow chromatography. The obtained endoglucanase ia a single subunit molecule about 31.5 kD or 34.5 kD as determined by 12% SDS-PAGE and gel filtration, respectively. The maximum endoglucanase activity was observed at pH 2.5 - 3.0 and 55℃. The endoglucanase appeared to be thermostable at 60 ℃, and exposure in 80℃ for 30 min resulted in 80% of activity loss. Metal ions like K^＋ , Ca^2＋ and Mn^2＋ enhanced the endoglucanase activity, whereas Ag^＋ , Cu^2＋ and Al^3＋ inhibited the activity significantly. The endoglucanase possessed a strong specificity to carboxymethyl cellulose.

关 键 词：嗜热子囊菌光孢变种 Thermoascus aurantiacus var.levisporus 内切β-葡聚糖酶 纯化 特性 

分 类 号：Q556.2[生物学—生物化学]