Yoon Jeong-Jun, Cha Chang-Jun, Kim Yeong-Suk, Son Dong-Won, Kim Young-Kyoon
Department ofBioscience and Biotechnology, Konkuk University, Seoul 143-701, Korea.
J Microbiol Biotechnol. 2007 May;17(5):800-5.
Two endoglucanases with processive cellulase activities, produced from Fomitopsis palustris grown on 2% microcrystalline cellulose (Avicel), were purified to homogeneity by anion-exchange and gel filtration column chromatography systems. SDS-PAGE analysis indicated that the molecular masses of the purified enzymes were 47 kDa and 35 kDa, respectively. The amino acid sequence analysis of the 47-kDa protein (EG47) showed a sequence similarity with fungal glycoside hydrolase family 5 endoglucanase from the white-rot fungus Phanerochaete chrysosporium. N-terminal and internal amino acid sequences of the 35-kDa protein (EG35), however, had no homology with any other glycosylhydrolases, although the enzyme had high specific activity against carboxymethyl cellulose, which is a typical substrate for endoglucanases. The initial rate of Avicel hydrolysis by EG35 was relatively fast for 48 h, and the amount of soluble reducing sugar released after 96 h was 100 microg/ml. Although EG47 also hydrolyzed Avicel, the hydrolysis rate was lower than that of EG35. Thin layer chromatography analysis of the hydrolysis products released from Avicel indicated that the main product was cellobiose, suggesting that the brown-rot fungus possesses processive EGs capable of degrading crystalline cellulose.
从生长在2%微晶纤维素（微晶纤维素）上的云芝中产生的两种具有连续纤维素酶活性的内切葡聚糖酶，通过阴离子交换和凝胶过滤柱色谱系统纯化至同质。SDS-PAGE分析表明，纯化酶的分子量分别为47 kDa和35 kDa。对47 kDa蛋白（EG47）的氨基酸序列分析显示，其与白腐真菌黄孢原毛平革菌的真菌糖苷水解酶家族5内切葡聚糖酶具有序列相似性。然而，35 kDa蛋白（EG35）的N端和内部氨基酸序列与任何其他糖基水解酶均无同源性，尽管该酶对羧甲基纤维素（内切葡聚糖酶的典型底物）具有高比活性。EG35对微晶纤维素的初始水解速率在48小时内相对较快，96小时后释放的可溶性还原糖量为100μg/ml。虽然EG47也能水解微晶纤维素，但其水解速率低于EG35。对微晶纤维素释放的水解产物进行薄层色谱分析表明，主要产物是纤维二糖，这表明褐腐真菌拥有能够降解结晶纤维素的连续内切葡聚糖酶。
