Improving the level of the cytidine biosynthesis in <i>E. coli</i> through atmospheric room temperature plasma mutagenesis and metabolic engineering
Xiangjun Zhang, Lu Liu, Cong Ma, Haojie Zhang, Huiyan Liu, Haitian Fang
Abstract
AIMS: Cytidine, as an important commercial precursor in the chemical synthesis of antiviral and antitumor drugs, is in great demand in the market. Therefore, the purpose of this study is to build a microbial cell factory with high cytidine production. METHODS AND RESULTS: A mutant E. coli NXBG-11-F34 with high tolerance to uridine monophosphate structural analogs and good genetic stability was obtained by atmospheric room temperature plasma (ARTP) mutagenesis combined with high-throughput screening. Then, the udk and rihA genes involved in cytidine catabolism were knocked out by CRISPR/Cas9 gene editing technology, and the recombinant strain E. coli NXBG-13 was constructed. The titer, yield, and productivity of cytidine fermented in a 5 l bioreactor were 15.7 g l-1, 0.164 g g-1, and 0.327 g l-1 h-1, respectively. Transcriptome analysis of the original strain and the recombinant strain E. coli NXBG-13 showed that the gene expression profiles of the two strains changed significantly, and the cytidine de novo pathway gene of the recombinant strain was up-regulated significantly. CONCLUSIONS: ARTP mutagenesis combined with metabolic engineering is an effective method to construct cytidine-producing strains.