Amino acids are key substrates to Escherichia coli BW25113 for achieving high specific growth rate

Kutscha

Pflügl

2020

Preprint

Background Acetate is an abundant carbon source and its use as an alternative feedstock has great potential for the production of fuel and platform chemicals. Acetoin and 2,3-butanediol represent two of these potential platform chemicals. Results The aim of this study was to produce 2,3-butanediol and acetoin from acetate in Escherichia coli W. The key strategies to achieve this goal were: strain engineering, in detail the deletion of mixed acid fermentation pathways E. coli W ΔldhA ΔadhE Δpta ΔfrdA 445_Ediss and the development of a new defined medium containing five amino acids and seven vitamins. Stepwise reduction of the media additives further revealed that diol production from acetate is mediated by the availability of aspartate. Other amino acids or TCA cycle intermediates did not enable growth on acetate. Cultivation under controlled conditions in batch and pulsed fed-batch experiments showed that aspartate was consumed before acetate, indicating that co-utilization is not a prerequisite for diol production. The addition of aspartate gave cultures a start-kick and was not required for feeding. Pulsed fed-batches resulted in the production of 1.43 g l-1 from aspartate and acetate and 1.16 g l-1 diols (2,3-butanediol and acetoin) from acetate alone. The yield reached 0.09 g diols per g acetate, which accounts for 26 % of the theoretical maximum.Conclusion This study for the first time showed acetoin and 2,3-butanediol production from acetate as well as the use of chemically defined medium for product formation from acetate in E. coli. Hereby, we provide a solid base for process intensification and the investigation of other potential products.

Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

Microbial upgrading of acetate into 2,3-butanediol and acetoin by E. coli W

Kutscha

Pflügl

2020

Preprint

“…When defined medium is used, all cellular components must be synthesized de novo, whereas different precursors can be metabolized when complex media components such as yeast extract are added. The drawback and limitations of the use of complex media components have recently been intensively investigated and described [20]. In short, utilization of yeast extract modulates growth parameters like the specific growth rate µ due to the depletion of components throughout the cultivation [20,21].…”

Section: Introductionmentioning

confidence: 99%

“…The drawback and limitations of the use of complex media components have recently been intensively investigated and described [20]. In short, utilization of yeast extract modulates growth parameters like the specific growth rate µ due to the depletion of components throughout the cultivation [20,21]. It leads to a lack in reproducibility due to variable composition [22] and to differences in cellular metabolism, e.g., in acetate excretion and protein expression [23].…”

Section: Introductionmentioning

confidence: 99%

Metabolic engineering of Escherichia coli W for isobutanol production on chemically defined medium and cheese whey as alternative raw material

Journal of Industrial Microbiology and Biotechnology

Baar

Freitag

et al. 2020

The aim of this study was to establish isobutanol production on chemically defined medium in Escherichia coli. By individually expressing each gene of the pathway, we constructed a plasmid library for isobutanol production. Strain screening on chemically defined medium showed successful production in the robust E. coli W strain, and expression vector IB 4 was selected as the most promising construct due to its high isobutanol yields and efficient substrate uptake. The investigation of different aeration strategies in combination with strain improvement and the implementation of a pulsed fed-batch were key for the development of an efficient production process. E. coli W ΔldhA ΔadhE Δpta ΔfrdA enabled aerobic isobutanol production at 38% of the theoretical maximum. Use of cheese whey as raw material resulted in longer process stability, which allowed production of 20 g l−1 isobutanol. Demonstrating isobutanol production on both chemically defined medium and a residual waste stream, this study provides valuable information for further development of industrially relevant isobutanol production processes.

“…Product formation from acetate has so far only been achieved by the addition of complex media components such as yeast extract. However, there are some major drawbacks and limitations related to the use of complex media additives [23]. It was recently shown that the availability of casamino acids led to differences in cellular metabolism, especially acetate metabolism in glucose-grown cultures [24].…”

mentioning

confidence: 99%

Microbial upgrading of acetate into 2,3-butanediol and acetoin by E. coli W

Kutscha

Pflügl

2020

Preprint

Background: Acetate is an abundant carbon source and its use as an alternative feedstock has great potential for the production of fuel and platform chemicals. Acetoin and 2,3-butanediol represent two of these potential platform chemicals.Results: The aim of this study was to produce 2,3-butanediol and acetoin from acetate in Escherichia coli W. The key strategies to achieve this goal were: strain engineering, in detail the deletion of mixed acid fermentation pathways E. coli W ΔldhA ΔadhE Δpta ΔfrdA 445_Ediss and the development of a new defined medium containing five amino acids and seven vitamins. Stepwise reduction of the media additives further revealed that diol production from acetate is mediated by the availability of aspartate. Other amino acids or TCA cycle intermediates did not enable growth on acetate. Cultivation under controlled conditions in batch and pulsed fed-batch experiments showed that aspartate was consumed before acetate, indicating that co-utilization is not a prerequisite for diol production. The addition of aspartate gave cultures a start-kick and was not required for feeding. Pulsed fed-batches resulted in the production of 1.43 g l-1 from aspartate and acetate and 1.16 g l-1 diols (2,3-butanediol and acetoin) from acetate alone. The yield reached 0.09 g diols per g acetate, which accounts for 26 % of the theoretical maximum.Conclusion: This study for the first time showed acetoin and 2,3-butanediol production from acetate as well as the use of chemically defined medium for product formation from acetate in E. coli. Hereby, we provide a solid base for process intensification and the investigation of other potential products.