Biochemical Engineering and Pilot Plant Research and Development Unit, King Mongkut's University of Technology Thonburi, 49 Soi Tientalay 25, Bangkhuntien-Chaytalay Rd., Thakhum, Bangkhuntien, Bangkok 10150, Thailand.
Bioprocess Biosyst Eng. 2013 Jan;36(1):45-56. doi: 10.1007/s00449-012-0760-y. Epub 2012 Jun 1.
We have constructed a reaction network model of Bacillus subtilis. The model was analyzed using a pathway analysis tool called elementary mode analysis (EMA). The analysis tool was used to study the network capabilities and the possible effects of altered culturing conditions on the production of a fibrinolytic enzyme, nattokinase (NK) by B. subtilis. Based on all existing metabolic pathways, the maximum theoretical yield for NK synthesis in B. subtilis under different substrates and oxygen availability was predicted and the optimal culturing condition for NK production was identified. To confirm model predictions, experiments were conducted by testing these culture conditions for their influence on NK activity. The optimal culturing conditions were then applied to batch fermentation, resulting in high NK activity. The EMA approach was also applied for engineering B. subtilis metabolism towards the most efficient pathway for NK synthesis by identifying target genes for deletion and overexpression that enable the cell to produce NK at the maximum theoretical yield. The consistency between experiments and model predictions proves the feasibility of EMA being used to rationally design culture conditions and genetic manipulations for the efficient production of desired products.
我们构建了枯草芽孢杆菌的反应网络模型。使用一种称为基本模式分析（EMA）的途径分析工具对模型进行了分析。该分析工具用于研究网络功能以及改变培养条件对枯草芽孢杆菌生产纤溶酶纳豆激酶（NK）的可能影响。基于所有现有的代谢途径，预测了在不同底物和氧气供应下 NK 在枯草芽孢杆菌中的最大理论产率，并确定了 NK 生产的最佳培养条件。为了验证模型预测，通过测试这些培养条件对 NK 活性的影响进行了实验。然后将最佳培养条件应用于分批发酵，从而获得高 NK 活性。还应用 EMA 方法对枯草芽孢杆菌的代谢进行工程改造，通过确定用于删除和过表达的靶基因，使细胞能够以最大理论产率生产 NK，从而找到最有效的 NK 合成途径。实验和模型预测之间的一致性证明了 EMA 用于合理设计培养条件和遗传操作以有效生产所需产品的可行性。
