A CRISPRi mediated self-inducible system for dynamic regulation of TCA cycle and improvement of itaconic acid production in Escherichia coli

Recent Progress in Metabolic Engineering of Escherichia coli for the Production of Various C4 and C5-Dicarboxylic Acids

“…Consequently, it may underline the need for an integrative, system-wide approach . With the aid of computational tools and genetically encoded biosensors, in silico metabolic analysis and adaptive laboratory evolution capable of genome-scale engineering have been applied in metabolic engineering. , These in silico tools and biosensors are also anticipated to reveal previously unknown enzymes with superior catalytic activity and enable more effective enzyme engineering than known enzymes. , They are expected to be also applied in DCAs production in E. coli . ,, …”

“…137,138 They are expected to be also applied in DCAs production in E. coli. 36,102,127 Furthermore, controls such as industrial-scale parameter studies can assist in optimizing culture conditions, potentially boosting DCA production in E. coli. 139 These achievements are expected to enable E. coli to have greater competitiveness for commercial mass production in more DCA cases, compared to natural DCA producers that currently struggle with economical production on a large scale.…”

“…ItcR attachment to its corresponding promoter, P ccl induces downstream expression of dCas9, suppressing the target gene with sgRNA. With CiMS, the transcription levels of icd , pykA , and sucCD genes were optimized, leading to a 1.23-fold rise in itaconate yield compared to the control strain, eventually generating 3.93 g/L of itaconate …”

“…A selfinducing system (CiMS) was developed using an itaconatesensing module comprised of an itaconate-responsive transcription factor, ItcR, from Yersinia pseudotuberculosis, and a flux interference module using CRISPRi. 102 ItcR attachment to its corresponding promoter, P ccl induces downstream expression of dCas9, suppressing the target gene with sgRNA. With CiMS, the transcription levels of icd, pykA, and sucCD genes were optimized, leading to a 1.23-fold rise in itaconate yield compared to the control strain, eventually generating 3.93 g/L of itaconate.…”

“…With CiMS, the transcription levels of icd, pykA, and sucCD genes were optimized, leading to a 1.23-fold rise in itaconate yield compared to the control strain, eventually generating 3.93 g/L of itaconate. 102 A succinate production strain with osmo-tolerant mutations was engineered to utilize an oxygen-sensitive bifunctional switch (OBS) to maintain efficient production under osmotic stress, potentially leading to decrease of production performance. 100 The OBS allowed the global regulator FNR to augment the expression of pck from Actinobacillus succinogenes under anaerobic conditions while reducing endogenous pykAF.…”