一种康氏木霉内切β-葡聚糖苷酶的分离纯化及酶学性质  

Purification of a new endo-β-glucanase from Trichoderma koningii and its characterization

作　　者：叶锦[1] 张静茹[1] 覃益民[1] 刘幽燕[1] 

机构地区：[1]广西大学化学化工学院,广西南宁530004

出　　处：《江苏农业学报》2015年第5期995-1000,共6页Jiangsu Journal of Agricultural Sciences

基　　金：国家自然科学基金项目(21276053);广西生物炼制重点实验室培育基地开放课题(GXBF11-04)

摘　　要：为获得康氏木霉(Trichodermakoningii)内切β-葡聚糖苷酶组分,并研究其酶学性质,通过硫酸铵盐析、DEAE Sepharose FF阴离子交换层析和Sephadex G-75凝胶过滤层析对康氏木霉GIMP3.444纤维素酶主要组分进行分离纯化,SDS-PAGE电泳检测蛋白质纯度和分子量,MALDI-TOFMS鉴定蛋白质种类。结果显示,从康氏木霉所产的纤维素酶系中分离纯化获得一种内切β-葡聚糖苷酶组分,相对分子质量为44 600,酶的比活力为19.65 U/mg。该内切酶的最适反应p H值为4.5,最适反应温度为55℃。以羧甲基纤维素钠(CMC-Na)为底物时,酶的米氏常数(Km)为7.22 mg/ml。不同金属离子对酶活性影响不同,其中Ca2+(0.3 mmol/L)对酶的抑制作用较强,抑制了近35%的活力。通过MALDI-TOFMS鉴定及数据库分析,确定该酶为一种新的内切β-葡聚糖苷酶组分。To identify the composition of cellulase produced by Trichodermakoningii, ammonium sulfate precipitati-on, DEAE-sepharose FF anion exchange chromatography and sephadex G-75 gel filtration chromatography were used for purification of the cellulase. The molecular weight and purity were determined by SDS-PAGE, and the protein was identi-fied by MALDI-TOFMS. A new endo-β-glucanase was identified with its molecular weight of 44 600 and the specific activity of 19. 65 U/mg. The optimum pH and temperature for the endo-β-glucanase were 4. 5 and 55 ℃. The Km value was 7. 22 mg/ml (CMC-Na). Ca2+(0. 3 mmol/L) could inhibit the enzymatic activity significantly by 35%. The enzymatic proper-ties of the new endo-β-glucanase was distinct from other endo-β-glucanase. It could be used to was analyze the synergetic mechanism of cellulases.

关 键 词：康氏木霉 内切Β-葡聚糖苷酶 分离纯化 酶学性质 

分 类 号：Q936[生物学—微生物学]