Influence of threonine exporters on threonine production in Escherichia coli (original) (raw)
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Threonine as a carbon source for Escherichia coli
Journal of bacteriology, 1981
Threonine can be used aerobically as the sole source of carbon and energy by mutants of Escherichia coli K-12. The pathway used involves the conversion of threonine via threonine dehydrogenase to aminoketobutyric acid, which is further metabolized by aminoketobutyric acid ligase, forming acetyl coenzyme A and glycine. A strain devoid of serine transhydroxymethylase uses this pathway and excretes glycine as a waste product. Aminoketobutyric acid ligase activity was demonstrated after passage of crude extracts through Sephadex G100.
Proteomic response analysis of a threonine-overproducing mutant of Escherichia coli
Biochemical Journal, 2004
The proteomic response of a threonine-overproducing mutant of Escherichia coli was quantitatively analysed by two-dimensional electrophoresis. Evidently, 12 metabolic enzymes that are involved in threonine biosynthesis showed a significant difference in intracellular protein level between the mutant and native strain. The level of malate dehydrogenase was more than 30-fold higher in the mutant strain, whereas the synthesis of citrate synthase seemed to be severely inhibited in the mutant. Therefore, in the mutant, it is probable that the conversion of oxaloacetate into citrate was severely inhibited, but the oxidation of malate to oxaloacetate was significantly up-regulated. Accumulation of oxaloacetate may direct the metabolic flow towards the biosynthetic route of aspartate, a key metabolic precursor of threonine. Synthesis of aspartase (aspartate ammonia-lyase) was significantly inhibited in the mutant strain, which might lead to the enhanced synthesis of threonine by avoiding un...
Biotechnology Progress, 2020
Phage infection is common during the production of L-threonine by E. coli, and low Lthreonine production and glucose conversion percentage are bottlenecks for the efficient commercial production of L-threonine. In this study, 20 antiphage mutants producing high concentration of Lthreonine were obtained by atmospheric and room temperature plasma (ARTP) mutagenesis, and an antiphage E. coli variant was characterized that exhibited the highest production of L-threonine (E. coli TRFC-AP). The elimination of fhuA expression in E. coli TRFC-AP was responsible for phage resistance. The biomass and cell growth of E. coli TRFC-AP showed no significant differences from those of the parent strain (E. coli TRFC), and the production of L-threonine (159.3 g L-1) and glucose conversion percentage (51.4%) were increased by 10.9% and 9.1%, respectively, compared with those of E. coli TRFC. During threonine production (culture time of 20 h), E. coli TRFC-AP exhibited higher activities of key enzymes for glucose utilization (HK, G6PD, PFK, PEPC, and PYK) and threonine synthesis (GS, AK, HSD, HSK and TS) compared to those of E. coli TRFC. The analysis of metabolic flux distribution indicated that the flux of threonine with E. coli TRFC-AP reached 69.8%, an increase of 16.0% compared with that of E. coli TRFC. Overall, higher L-threonine production and glucose conversion percentage were obtained with E. coli TRFC-AP due to increased activities of key enzymes and improved carbon flux for threonine synthesis.
A study on L-threonine and L-serine uptake in Escherichia coli K-12
Frontiers in Microbiology
In the current study, we report the identification and characterization of the yifK gene product as a novel amino acid carrier in E. coli K-12 cells. Both phenotypic and biochemical analyses showed that YifK acts as a permease specific to L-threonine and, to a lesser extent, L-serine. An assay of the effect of uncouplers and composition of the reaction medium on the transport activity indicates that YifK utilizes a proton motive force to energize substrate uptake. To identify the remaining threonine carriers, we screened a genomic library prepared from the yifK-mutant strain and found that brnQ acts as a multicopy suppressor of the threonine transport defect caused by yifK disruption. Our results indicate that BrnQ is directly involved in threonine uptake as a low-affinity but high-flux transporter, which forms the main entry point when the threonine concentration in the external environment reaches a toxic level. By abolishing YifK and BrnQ activity, we unmasked and quantified the ...
Threonine analogue resistant mutants of Escherichia coli K-12
Biotechnology Letters, 1995
Mutants of Escherichia coli K-12 resistant to a threonine analogue (ot-amino-I]-hydroxy valeric acid) were predominantly resistant to ethionine and overproduced both threonine and methionine (2 mg/ml each). Novelty of the mutants is discussed.
A novel membrane-associated threonine permease encoded by the tdcC gene of Escherichia coli
Journal of bacteriology, 1990
A novel L-threonine transport system is induced in Escherichia coli cells when incubated in amino acid-rich medium under anaerobic conditions. Genetic and biochemical analyses with plasmids harboring mutations in the anaerobically expressed tdcABC operon indicated that the tdcC gene product was responsible for L-threonine uptake. Competition experiments revealed that the L-threonine transport system is also involved in L-serine uptake and is partially shared for L-leucine transport; L-alanine, L-valine, and L-isoleucine did not affect L-threonine uptake. Transport of L-threonine was inhibited by the respiratory chain inhibitors KCN and carbonyl cyanide m-chlorophenylhydrazone and was Na+ independent. These results identify for the first time an E. coli gene encoding a permease specific for L-threonine-L-serine transport that is distinct from the previously described threonine-serine transport systems. A two-dimensional topological model predicted from the amino acid composition and ...
Role of threonine dehydrogenase in Escherichia coli threonine degradation
Journal of bacteriology, 1977
Threonine was used as nitrogen source by Escherichia coli K-12 through a pathway beginning with the enzyme threonine dehydrogenase. The 2-amino-3-ketobutyrate formed was converted to glycine, and the glycine was converted to serine, which acted as the actual nitrogen donor. The enzyme formed under anaerobic conditions and known as threonine deaminase (biodegradative) is less widespread than threonine dehydrogenase and may be involved in energy metabolism rather than in threonine degradation per se.
2002
The industrial production of compounds by E. coli strains is often accompanied with variability in yield levels. To investigate the mechanism of such instability the over-production of threonine was used as a model. The instability in this strain appears to be caused by a metabolic burden resulting in occurrence of low producing revertants. A successful application of the tightly regulated T7 expression system is presented as a possible solution providing a substantial stabilization of the threonine production.
2018
Department of Physiology, Vidyasagar College, 39, Sankar Ghosh Lane, Kolkata-700 006, India <em>E</em>-<em>mail:</em> res_biol@rediffmail.com Department of Chemical Engineering, Biochemical Engineering Division, Biotechnology Laboratory, University of Calcutta, 92, Acharya Prafulla Chandra Road, Kolkata-700 009, India P.G. Department of Botany, Utkal University, Vani Vihar, Bhubaneswar-751 004, Odisha, India <em>Manuscript received 16 July 2018, accepted 09 August 2018</em> A high yielding L-threonine auxotrophic mutant <em>Corynebacterium glutamicum</em> X1870 was developed by induced mutation using UV irradiations as a physical mutagen followed by penicillin selection. It was an a,E-diamopimilic acid (DAP) deficient mutant. Production of L-threonine was greatly influenced by thiamine-HCl (60 µg/L) and DAP (60 mg/L) in the synthetic medium with D-glucose as a carbon source at 30°C after 72 h of incubation. The production of L-threon...