Key Laboratory of Systems Microbial Biotechnology, Tianjin Institute of Industrial Biotechnology, Chinese Academy of Sciences, Tianjin 300308, China.
Key Laboratory of Synthetic Biology, Institute of Plant Physiology and Ecology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200032, China.
Metab Eng. 2017 Nov;44:13-21. doi: 10.1016/j.ymben.2017.08.005. Epub 2017 Aug 31.
The MEP pathway genes were modulated to investigate whether there were new rate-limiting steps and toxic intermediates in this pathway. Activating IspG led to significant decrease of cell growth and β-carotene production. It was found that ispG overexpression led to accumulation of intermediate HMBPP, which seriously interfered with synthesis machinery of nucleotide and protein in Escherichia coli. Activation of the downstream enzyme IspH could solve HMBPP accumulation problem and eliminate the negative effects of ispG overexpression. In addition, intermediate MECPP accumulated in the starting strain, while balanced activation of IspG and IspH could push the carbon flux away from MECPP and led to 73% and 77% increase of β-carotene and lycopene titer respectively. Our work for the first time identified HMBPP to be a cytotoxic intermediate in MEP pathway and demonstrated that balanced activation of IspG and IspH could eliminate accumulation of HMBPP and MECPP and improve isoprenoids production.
对 MEP 途径基因进行了调节，以研究该途径是否存在新的限速步骤和有毒中间产物。激活 IspG 导致细胞生长和 β-胡萝卜素产量显著下降。研究发现，ispG 的过表达导致中间产物 HMBPP 的积累，这严重干扰了大肠杆菌核苷酸和蛋白质合成机制。激活下游酶 IspH 可以解决 HMBPP 积累问题，并消除 ispG 过表达的负面影响。此外，中间产物 MECPP 在起始菌株中积累，而平衡激活 IspG 和 IspH 可以将碳通量从 MECPP 中转移，分别使 β-胡萝卜素和番茄红素的产量增加 73%和 77%。我们的工作首次确定 HMBPP 是 MEP 途径中的细胞毒性中间产物，并证明平衡激活 IspG 和 IspH 可以消除 HMBPP 和 MECPP 的积累，提高异戊二烯产量。
