The Effect of E. coli Uridine-Cytidine Kinase Gene Deletion on Cytidine Synthesis and Transcriptome Analysis

Abstract

Cytidine is an antiviral and anticancer drug intermediate, its primary method of manufacture being fermentation. Uridine-cytidine kinase (UCK) catalyzes the reverse process of phosphorylation of cytidine to produce cytidylic acid, which influences cytidine accumulation in the Escherichia coli cytidine biosynthesis pathway. The cytidine-producing strain E. coli NXBG-11 was used as the starting strain in this work; the udk gene coding UCK was knocked out of the chromosomal genome using clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology. The mutant strain E. coli NXBG-12 was obtained; its transcriptomics were studied to see how udk gene deletion affected cytidine synthesis and cell-wide transcription. The mutant strain E. coli NXBG-12 generated 1.28 times more cytidine than the original strain E. coli NXBG-11 after 40 h of shake-flask fermentation at 37 °C. The udk gene was knocked out, and transcriptome analysis showed that there were 1168 differentially expressed genes between the mutant and original strains, 559 upregulated genes and 609 downregulated genes. It was primarily shown that udk gene knockout has a positive impact on the cytidine synthesis network because genes involved in cytidine synthesis were significantly upregulated (p < 0.05) and genes related to the cytidine precursor PRPP and cofactor NADPH were upregulated in the PPP and TCA pathways. These results principally demonstrate that udk gene deletion has a favorable impact on the cytidine synthesis network. The continual improvement of cytidine synthesis and metasynthesis is made possible by this information, which is also useful for further converting microorganisms that produce cytidine.