Department of Chemical and Biological Engineering, South Dakota School of Mines and Technology, Rapid City, SD 57701, USA.
Bioresour Technol. 2010 Nov;101(22):8798-806. doi: 10.1016/j.biortech.2010.06.001.
The composition of thermophilic (60 degrees C) mixed cellulose-degrading enrichment culture initiated from compost samples was examined by constructing a 16S rRNA gene clone library and the presence of sequences related to Actinobacteria, Bacteroidetes, Chloroflexi, Deinococcus-Thermus, Firmicutes, and Proteobacteria were identified. Eight isolates capable of degrading cellulose, carboxymethyl cellulose (CMC), or ponderosa pine sawdust were identified as belonging to the genera Geobacillus, Thermobacillus, Cohnella, and Thermus. A compost isolate WSUCF1 (Geobacillus sp.) was selected based on its higher growth rate and cellulase activity compared to others in liquid minimal medium containing cellulose as a source of carbon and energy. Strain WSUCF1 and a previously isolated thermophilic cellulose-degrading deep gold mine strain DUSELR13 (Bacillus sp.) were examined for their enzyme properties and kinetics. The optimal pH for carboxymethyl cellulase (CMCase) activity was 5.0 for both isolates. The optimum temperatures for CMCase of WSUCFI and DUSELR13 were 70 and 75 degrees C, respectively. For CMC, the DUSELR13 and WSUCF1 CMCases had K(m) values of 3.11 and 1.08mg/ml, respectively. Most remarkably, WSUCF1 and DUSELR13 retained 89% and 78% of the initial CMCase activities, respectively, after incubation at 70 degrees C for 1day. These thermostable enzymes would facilitate development of more efficient and cost-effective forms of the simultaneous saccharification and fermentation process to convert lignocellulosic biomass into biofuels.
从堆肥样本中启动的嗜热（60°C）混合纤维素降解富集培养物的组成，通过构建 16S rRNA 基因克隆文库进行了检查，鉴定出与放线菌、拟杆菌门、绿弯菌门、厚壁菌门、变形菌门相关的序列。鉴定出 8 个能够降解纤维素、羧甲基纤维素（CMC）或辐射松锯末的分离株，属于芽孢杆菌属、芽孢杆菌属、科恩氏菌属和嗜热菌属。基于其在含有纤维素作为碳源和能源的液体最小培养基中的生长速度和纤维素酶活性高于其他菌株，选择堆肥分离株 WSUCF1（芽孢杆菌属）。对 WSUCF1 菌株和先前分离的嗜热纤维素降解深层金矿菌株 DUSELR13（芽孢杆菌属）进行了酶特性和动力学研究。两种分离株的羧甲基纤维素酶（CMCase）活性的最佳 pH 值均为 5.0。WSUCF1 和 DUSELR13 的 CMCase 最适温度分别为 70 和 75°C。对于 CMC，DUSELR13 和 WSUCF1 的 CMCases 的 K(m) 值分别为 3.11 和 1.08mg/ml。值得注意的是，WSUCF1 和 DUSELR13 在 70°C 孵育 1 天后，分别保持初始 CMCase 活性的 89%和 78%。这些耐热酶将有助于开发更高效、更具成本效益的同步糖化和发酵形式，将木质纤维素生物质转化为生物燃料。
