Ameliorating end-product inhibition to improve cadaverine production in engineered Escherichia coli and its application in the synthesis of bio-based diisocyanates

Abstract

Cadaverine is an important C5 platform chemical with a wide range of industrial applications. However, the cadaverine inhibition on the fermenting strain limited its industrial efficiency of the strain. In this study, we report an engineered Escherichia coli strain with high cadaverine productivity that was generated by developing a robust host coupled with metabolic engineering to mitigate cadaverine inhibition. First, a lysine producing E. coli was treated with a combination of radiation (ultraviolet and visible spectrum) and ARTP (atmospheric and room temperature plasma) mutagenesis to obtain a robust host with high cadaverine tolerance. Three mutant targets including HokD, PhnI and PuuR are identified for improved cadaverine tolerance. Further transcriptome analysis suggested that cadaverine suppressed the synthesis of ATP and lysine precursor. Accordingly, the related genes involved in glycolysis and lysine precursor, as well as cadaverine exporter was engineered to release the cadaverine inhibition. The final engineered strain was fed-batch cultured and a titer of 58.7 g/L cadaverine was achieved with a yield of 0.396 g/g, both of which were the highest level reported to date in E. coli . The bio-based cadaverine was purified to >99.6% purity, and successfully used for the synthesis of polyurethane precursor 1,5-pentamethylene diisocyanate (PDI) through the approach of carbamate decomposition.

A novel co-production of cadaverine and succinic acid based on a thermal switch system in recombinant Escherichia coli

“…To construct succinic acid producer strain, the genes of ldh and ackA were knocked out from E. coli NT1003 by CRISPR-cas9 system. The procedures were described as Xin Wang et al [ 6 ]. E. coli trans1 T1 used to construct recombinant plasmids was cultured in Luria–Bertani (LB) medium, consisting of 10 g/L peptone, 5 g/L yeast extract, 5 g/L NaCl.…”

“…The biomass was monitored by measuring OD 600 . The concentration of lysine and glucose was measure by SBA biosensor [ 6 ]. The cadaverine and succinate acid was analyzed by HPLC [ 6 , 16 ].…”

“…In procaryotic organism system, cadaverine is synthesized through the glycolysis pathway, tricarboxylic acid cycle, L-lysine synthesis pathway and DAP pathway with renewable carbon sources like glucose as substrate [ 4 ]. To enhance production of cadaverine, multiple metabolic engineering strategies were researched and employed into improve metabolic flux towards lysine, decrease synthesis of byproducts and enhance supply of cofactor [ 5 , 6 ]. However, these metabolic modification often led to unbalanced cellular metabolic network, resulting in inhibition of cell growth and restriction of production [ 7 ].…”

Advances and prospects in the biosynthesis of vitamin B6 and its bioengineering as a cofactor of other chemicals

“…Compared with the traditional petrochemical‐based polyamides, the biological polyamide with cadaverine has excellent physical properties (polyamides 6 and 66) (Leong et al, 2020). It has become a research hotspot in recent years and some researchers are committed to using metabolic engineering strategies to improve its production (Imao et al, 2017; Ma et al, 2015; Wang et al, 2021). In recent years, various PLP‐dependent enzymes have been developed for industrial applications, especially for whole‐cell biocatalysis and the proper function of enzymes also depends on sufficient levels of PLP in cells (Schiroli & Peracchi, 2015; Steffen‐Munsberg et al, 2015).…”

Separation, purification, and crystallization of 1,5-pentanediamine hydrochloride from fermentation broth by cation resin

“…At present, the industrial preparation of PDI is mainly liquid-phase phosgenation ( Li W et al, 2021 ). The raw material for PDI is 1,5-pentanediamine (PDA), which needs to be obtained from biomass such as feed corn starch through biological fermentation engineering ( Wang et al, 2021 ). In the two-step reaction, PDA initially reacts with a cold phosgene solution to produce 1,5-pentanediamine hydrochloride (PDAH) and carbamate.…”