Kaur Jatinder, Chadha Bhupinder S, Kumar Badhan A, Saini Harvinder S
Department of Microbiology, Guru Nanak Dev University, Amritsar 143 005, India.
Bioresour Technol. 2007 Jan;98(1):74-81. doi: 10.1016/j.biortech.2005.11.019. Epub 2006 Jan 10.
This study reports the purification and characterization of endoglucanases (EG I and EG II) from a newly isolated thermophilic fungus, Melanocarpus sp. MTCC 3922. The molecular weight of EG I and EG II as with SDS-PAGE and pI were approximately 40 and 50 kDa, and approximately 4.0 and 3.6, respectively. EG I and EG II were optimally active at 50 and 70 degrees C, and pH 6.0 and 5.0, respectively. EG I was active over a broad range of pH (5.0-7.0), whereas, loss of activity was observed as the temperature was increased from 50 to 80 degrees C. However, EG II was active over pH 4.0-6.0 and temperature 40-80 degrees C. The presence of mercaptoethanol and SDS inhibited the EG I activity but showed no negative effect on EG II. Both the endoglucanases showed higher activity against barley-beta-glucan as compared to CMC. Km values of EG I and EG II for barley-beta-glucan were lower than CMC. Turn over number (K(cat)) and catalytic efficiency (K(cat)/Km) values of both the endoglucanases were higher with barley-beta-glucan as substrate than CMC. EG I showed affinity for Avicel indicating the presence of cellulose binding domains (CBD) whereas, EG II was found to lack CBD.
本研究报道了从新分离的嗜热真菌黑果菌属MTCC 3922中纯化和鉴定内切葡聚糖酶（EG I和EG II）的过程。通过SDS-PAGE测定，EG I和EG II的分子量分别约为40 kDa和50 kDa，其等电点分别约为4.0和3.6。EG I和EG II的最适活性温度分别为50℃和70℃，最适pH分别为6.0和5.0。EG I在较宽的pH范围（5.0 - 7.0）内具有活性，然而，当温度从50℃升高到80℃时，其活性丧失。而EG II在pH 4.0 - 6.0和温度40 - 80℃范围内具有活性。巯基乙醇和SDS的存在会抑制EG I的活性，但对EG II没有负面影响。与羧甲基纤维素（CMC）相比，这两种内切葡聚糖酶对大麦β-葡聚糖均表现出更高的活性。EG I和EG II对大麦β-葡聚糖的米氏常数（Km）低于CMC。以大麦β-葡聚糖为底物时，这两种内切葡聚糖酶的转换数（K(cat)）和催化效率（K(cat)/Km）均高于以CMC为底物时。EG I对微晶纤维素有亲和力，表明存在纤维素结合结构域（CBD），而EG II则缺乏CBD。
