State Key Laboratory of Microbial Technology, National Glycoengineering Research Center , Shandong University , Qingdao 266237 , P. R. China.
CAS Key Lab of Biobased Materials, Qingdao Institute of Bioenergy and Bioprocess Technology , Chinese Academy of Sciences , Qingdao 266101 , P. R. China.
J Agric Food Chem. 2019 Feb 6;67(5):1478-1483. doi: 10.1021/acs.jafc.8b06496. Epub 2019 Jan 25.
5-Aminolevulinic acid (5-ALA) is a key metabolic intermediate of the heme biosynthesis pathway, which has broad application prospects in agriculture and medicine. However, segregational instability of plasmid-based expression systems and low yield have hampered large-scale manufacture of 5-ALA. In this study, two important genes of the 5-ALA C5 biosynthesis pathway, hemA and hemL, were integrated into Escherichia coli MG1655 for chemically induced chromosomal evolution (CIChE). The highest hemA and hemL copy-number, 98 per genome, was obtained in CIChE strain MG136. The 5-ALA titer of this strain reached 2724 mg/L in optimized condition. Then, after undergoing adaptative evolution and the deletion of recA, strain MG136a ΔrecA::FRT could stably produce 4550 mg/L 5-ALA from glucose, 450 times the amount produced by hemA- hemL single copy strain MG1655-hemAL. This study constructed a plasmid-free E. coli strain for 5-ALA production, which will provide the basis for further manipulation of metabolic regulation and optimization of fermentation.
5-氨基酮戊酸（5-ALA）是血红素生物合成途径的关键代谢中间产物，在农业和医学中有广泛的应用前景。然而，基于质粒的表达系统的分离不稳定性和低产量阻碍了 5-ALA 的大规模生产。在本研究中，将 5-ALA C5 生物合成途径的两个重要基因 hemA 和 hemL 整合到大肠杆菌 MG1655 中，进行化学诱导染色体进化（CIChE）。在 CIChE 菌株 MG136 中获得了最高的 hemA 和 hemL 拷贝数，每个基因组 98 个。在优化条件下，该菌株的 5-ALA 产量达到 2724mg/L。然后，经过适应性进化和 recA 的缺失，菌株 MG136a ΔrecA::FRT 可以从葡萄糖稳定地生产 4550mg/L 的 5-ALA，是单拷贝菌株 MG1655-hemAL 的 450 倍。本研究构建了无质粒的大肠杆菌菌株用于 5-ALA 的生产，这将为进一步进行代谢调控和发酵优化提供基础。
