ACTIVITY STUDIES OF 8 PURIFIED CELLULASES - SPECIFICITY, SYNERGISM, AND BINDING DOMAIN EFFECTS
ACTIVITY STUDIES OF 8 PURIFIED CELLULASES - SPECIFICITY, SYNERGISM, AND BINDING DOMAIN EFFECTS
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DOI：
10.1002/bit.260420811
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发表时间：
1993-10-01
影响因子：
3.8
通讯作者：
WILSON, DB
中科院分区：
文献类型：
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作者：
IRWIN, DC;SPEZIO, M;WILSON, DB
分别在滤纸、膨化纤维素和CMC上测定了纯化的6种褐热孢子菌纤维素酶和里氏木霉CBHI和CBHII的活性。发现了一种测定滤纸水解产物中可溶性和不可溶性还原糖的简便方法，可以有效地区分胞外酶和胞内酶。胞内酶产生34%至50%的不溶性还原糖，胞外酶产生不到8%的不溶性还原糖。这些纤维素酶的多种混合物在16小时内消化5.2%滤纸的能力进行了定量测量。混合酶的比活性为每微摩尔酶每分钟0.41 ~ 16.31 μ mol纤维素糖。协同作用程度为0.4 ~ 7.8。发现T. reesei CBHII和T. fusca E3在混合物中功能相当。纯化了褐霉内胞酶E2和E5的催化结构域（cd），发现它们分别保持了93%和100%的CMC活性，但cd蛋白都不能消化滤纸达到5.2%。当E2cd和E5cd以协同混合物取代天然蛋白时，含有E2cd的混合物保留了60%的活性，含有E5cd的混合物保留了94%的活性。添加β -葡萄糖苷酶可使最佳增效混合物的活性增加一倍。枸杞粗纤维素酶在滤纸上的活性增加了1.7倍。1993 John Wiley & Sons, Inc.；
The activities of six purified Thermomonospora fusca cellulases and Trichoderma reesei CBHI and CBHII were determined on filter paper, swollen cellulose, and CMC. A simple method to measure the soluble and insoluble reducing sugar products from the hydrolysis of filter paper was found to effectively distinguish between exocellulases and endocellulases. Endocellulases produced 34% to 50% insoluble reducing sugar and exocellulases produced less than 8% insoluble reducing sugar. The ability of a wide variety of mixtures of these cellulases to digest 5.2% of a filter paper disc in 16 h was measured quantitatively. The specific activities of the mixtures varied from 0.41 to 16.31 mumol cellobiose per minute per micromole enzyme. The degree of synergism ranged from 0.4 to 7.8. T. reesei CBHII and T. fusca E3 were found to be functionally equivalent in mixtures. The catalytic domains (cd) of T. fusca endocellulases E2 and E5 were purified and found to retain 93% and 100% of their CMC activity, respectively, but neither cd protein could digest filter paper to 5.2%. When E2cd and E5cd were substituted in synergistic mixtures for the native proteins, the mixtures containing E2cd retained 60%, and those containing E5cd retained 94% of the original activity. Addition of a beta-glucosidase was found to double the activity of the best synergistic mixture. Addition of CBHI to T. fusca crude cellulase increased its activity on filter paper 1.7-fold. (C) 1993 John Wiley & Sons, Inc.