研究报告
张耿崚,韩业钜,陈细妹,王小琴,约比,黄佳钦,叶志超,李清荷.产纤维素酶嗜热地芽孢杆菌HTA426的筛选鉴定、酶学性质分析及其应用[J].环境科学学报,2017,37(4):1444-1453
产纤维素酶嗜热地芽孢杆菌HTA426的筛选鉴定、酶学性质分析及其应用
- Isolation, identiafication, characterization and application of cellulase-producing Geobacillus kaustophilus HTA426
- 基金项目：广东工业大学青年基金（No.14QNZD005）；广东工业大学青年百人A类项目（No.1143-220413105）；广东省教育厅高等院校学科建设专项资金项目（No.2014KTSP022）：广东工业大学大学生创新创业训练计划项目（No.xj201511845089，yj201611845133）
- 作者
- 单位
- 张耿崚
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 韩业钜
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 陈细妹
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 王小琴
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 约比
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 黄佳钦
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 叶志超
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 李清荷
- 广东工业大学环境科学与工程学院和环境健康与污染控制研究院, 广州 510006
- 摘要：为解决现今纤维素酶生产的低效率和高成本问题，以及实现利用木质纤维素废弃物生产纤维素酶的目的，本研究结合羧甲基纤维素钠（CMC）水解圈法和胞外酶活测定法，从嗜热菌资源丰富的温泉区土壤中筛选出一株具有产羧甲基纤维素酶（CMCase）的嗜热地芽孢杆菌HTA426（Geobacillus kaustophilus HTA426）.对菌株生产的酶进行酶学性质分析并将菌株应用于以木质纤维素为替代碳源产纤维素酶.采用硫酸铵沉淀法和离子交换层析法对粗酶液进行二级纯化，回收率和纯化倍数分别为10.14%和5.12，纯化后的CMCase的分子量约为40 kDa.其CMCase活性在温度为60℃，pH为7.0下达到最高.培育5 h后，酶的活性在温度为50~70℃下也能保持相对稳定.HAT426菌株能够在已经经过碱处理的甘蔗渣、稻杆及水葫芦为碳源的培养液中生长和生产CMCase.以甘蔗渣为碳源的培养液最适合于生产CMCase，其活性为103.67 U·mL-1.菌株HTA426是首次报道的具有产纤维素酶能力的嗜热地芽孢杆菌，经纯化的纤维素酶具有广泛的pH适用性（pH 5~8）和良好的热稳定性（50~70℃），在运用于木质纤维素能源化中具有较大潜力.
- Abstract：To reduce the high cost and increase the productivity of cellulase from lignocellulose waste, a bacterial strain (HTA426) producing thermophilic carboxymethyl cellulase (CMCase) was isolated from hot spring in this study, and was identified as Geobacillus kaustophilus. The enzymatic properties of cellulase produced by the bacteria were investigated and then the strain also was applied for cellulase production using the lignocellulose as alternative carbon source. The two-step purification of crude enzyme was carried out by ammonium sulfate precipitation and ion exchange chromatography, the recovery and purification fold were 10.14% and 5.12, respectively. The molecular mass of the purified CMCase was 40 kDa. The enzyme had the highest CMCase activity at 60 ℃ and pH 7.0. It was also found that the enzyme kept stable over a broad temperature range of 50~70 ℃ after 5 h incubation. Moreover, strain HTA426 was able to grow and produce CMCase on the medium supplemented with alkali-treated sugarcane bagasse, rice straw and waterhyacinth as carbon sources. Sugarcane bagasse was the best carbon source for the CMCase production with a CMCase activity of 103.67 U·mL-1. The strain HTA426 was a novel reported cellulase producing bacterium from Geobacillus kaustophilus. Due to its extensive pH adaptation (pH 5~8) and excellent temperature stability (50~70 ℃), the purified enzyme from this strain has a great potential in recovery of lignocellulose biomass waste into biofuel.
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