Control of fed-batch fermentations

Lee, Jeong-Seok; Lee, Sang Yup; Park, Sun Won; Middelberg, Anton P. J.

doi:10.1016/s0734-9750(98)00015-9

Cited by 251 publications

(161 citation statements)

References 49 publications

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“…In order to increase pyruvate yield from glucose, cell growth must be limited and excess glucose supplied. An exponential fed-batch process is ideal to limit growth at a constant rate, reducing the aeration demand and offering controlled growth conditions (4,21). We selected the unoptimized growth rate of 0.15 h Ϫ1 to focus our study of the effects of nutrient limitation and genetic perturbations on glycolytic flux and pyruvate yield.…”

Section: Discussionmentioning

confidence: 99%

High Glycolytic Flux Improves Pyruvate Production by a Metabolically Engineered Escherichia coli Strain

Zhu

Eiteman

Altman

et al. 2008

Appl Environ Microbiol

105

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We report pyruvate formation in Escherichia coli strain ALS929 containing mutations in the aceEF, pfl, poxB, pps, and ldhA genes which encode, respectively, the pyruvate dehydrogenase complex, pyruvate formate lyase, pyruvate oxidase, phosphoenolpyruvate synthase, and lactate dehydrogenase. The glycolytic rate and pyruvate productivity were compared using glucose-, acetate-, nitrogen-, or phosphorus-limited chemostats at a growth rate of 0.15 h ؊1 . Of these four nutrient limitation conditions, growth under acetate limitation resulted in the highest glycolytic flux (1.60 g/g · h), pyruvate formation rate (1.11 g/g · h), and pyruvate yield (0.70 g/g). Additional mutations in atpFH and arcA (strain ALS1059) further elevated the steady-state glycolytic flux to 2.38 g/g · h in an acetate-limited chemostat, with heterologous NADH oxidase expression causing only modest additional improvement. A fed-batch process with strain ALS1059 using defined medium with 5 mM betaine as osmoprotectant and an exponential feeding rate of 0.15 h ؊1 achieved 90 g/liter pyruvate, with an overall productivity of 2.1 g/liter · h and yield of 0.68 g/g.Pyruvic acid (pyruvate) is widely used in food, chemicals, and pharmaceuticals. The chemical is a precursor for the enzymatic production of L-tryptophan, L-tyrosine, D-/L-alanine, and L-dihydroxyphenylalanine (22), and it also serves in several health-related roles, including weight loss (25,33,34), exercise endurance (32), cholesterol reduction (35), and acne treatment (10). Recently, pyruvate has been used as the key metabolic precursor to the second-generation biofuels isobutanol and 3-methyl-1-butanol (2). By applying metabolic engineering strategies to them, microorganisms such as Escherichia coli and yeasts can be used to produce significant quantities of pyruvate from glucose and other renewable resources (22). In general, such approaches must delete or repress pathways which metabolize pyruvate. For example, pyruvate accumulates readily in E. coli strains having mutations in aceEF, encoding components of the pyruvate dehydrogenase complex (37). Additional mutations, of the ldhA, poxB, pfl, and pps genes, further improve pyruvate formation (48). Figure 1 shows the principal metabolic pathways and enzymes involved in the formation of pyruvate.Because pyruvate resides biochemically at the end of glycolysis, pyruvate production is directly related to the glycolytic flux. Metabolic engineering strategies to form pyruvate therefore also aim to enhance glycolysis (8,45,46). Glycolysis is not transcriptionally limited, and control principally resides outside the pathway in cellular demand for global cofactors, such as ATP and NADH (19,23,40). Glycolytic flux is substantially increased by disrupting oxidative phosphorylation or by increasing ATP hydrolysis (8,19). Increased glycolytic flux assists pyruvate accumulation: for example, an F 1 -ATPase-defective mutant (E. coli lipA2 bgl ϩ atpA401) generated pyruvate more quickly than its parent (45,46). Similarly, E. coli atpFH (strain TC44), defi...

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Section: Discussionmentioning

confidence: 99%

High Glycolytic Flux Improves Pyruvate Production by a Metabolically Engineered Escherichia coli Strain

Zhu

Eiteman

Altman

et al. 2008

Appl Environ Microbiol

105

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“…Fed-batch is widely used in lab-scale protein production. 14 Fed-batch offers the following advantages: 1) feeding rate is a good way to control the microorganism's growth according to its physiology, such as to reach a high cell density, 2) fermentation optimization is an important step in preventing by-product formation 15 and inhibition of substrate-associated growth, 16 and 3) productive phase is extendable under controlled conditions, giving high yield. The parameters of fed-batch involved substrate level, temperature, and pH, and these parameters are affected by each other.…”

Section: Introductionmentioning

confidence: 99%

High purity recombinant human growth hormone (rhGH) expression in Escherichia coli under phoA promoter

et al. 2016

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ABSTACTRecombinant human Growth Hormone (rhGH) is an important protein for human growth and is in high demand in clinics. Hence, it is necessary to develop an efficient fermentation process to produce highly pure rhGH. In this study, rhGH was expressed in Escherichia coli under alkaline phosphatase (phoA) promoter. The cultivation conditions for high expression level and purity of rhGH were investigated. The best initial phosphate concentration for rhGH expression, out of the 4 levels of initial phosphate concentration tests performed, was 12.6 mmol/L. Subsequently, 2 fed-batch cultivations under low dissolved oxygen (DO) (0% -10%) and high DO (20% -30%) conditions were carried out. High purity rhGH (92%) was obtained from 20% -30% DO-stat cultivation, although the biomass did not show any significant difference. In summary, this research provided an efficient fermentation process for high purity rhGH production from E. coli under phoA promoter, which can lower the production and purification costs for large-scale production of rhGH.

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“…Já o processo descontínuo alimentado de cultivo implica na adição de um ou mais nutrientes no decorrer do processo, possibilitando desta maneira a prevenção de efeitos inibitórios ou tóxicos por parte de determinados nutrientes, e até mesmo o deslocamento do metabolismo celular para a obtenção de um determinado produto (LEE et al, 1999;SCHMIDELL et al, 2001). No caso de cultivos de A. platensis, a utilização de fontes de nitrogênio que levam à formação de amônia no meio de cultivo faz com que o processo descontínuo alimentado seja necessário (BEZERRA, 2006;FERREIRA, 2008), pois tal nutriente, quando em altas concentrações, pode ser inibitório ou tóxico à cianobactéria (ABELIOVICH; AZOV, 1976;YUAN et al, 2011).…”

Section: Processos De Cultivounclassified

Avaliação da utilização do dióxido de carbono proveniente da fermentação alcoólica no cultivo de Spirulina (Arthrospira) platensis: uso simultâneo de nitrato de sódio e sulfato de amônio como fontes de nitrogênio em fotobiorreator aberto

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O principal objetivo deste trabalho foi a avaliação do potencial da utilização do dióxido de carbono proveniente da fermentação alcoólica no cultivo Spirulina (Arthrospira) platensis, visando demonstrar a possibilidade do uso de um gás efluente na produção de biomassa microbiana de alto valor comercial. Para tanto, tal cianobactéria foi cultivada em tanques abertos, em escala laboratorial, em temperatura de 30 ± 1 °C e intensidade luminosa de 156 ± 20 µmol fótons m -2 s -1 . O estudo de diversas variáveis de cultivo levou à fixação das seguintes condições: concentração do inóculo de 400 ± 20 mg L -1 ; pH de 9,0 ± 0,3, controlado por meio da adição de dióxido de carbono proveniente de cilindros; meio de cultura Schlösser, modificado de maneira a conter 0,497 e 16,4 g L -1 de carbonato e bicarbonato de sódio, respectivamente, e apenas 5,9 mM de nitrato de sódio; adição de 7,5 mM de sulfato de amônio no decorrer de 13 dias, em quantidades diárias exponencialmente crescentes, através do processo descontínuo alimentado de cultivo. Sob tais condições foram obtidos os seguintes resultados: concentração celular máxima (X m ) de 2990 mg L -1 , produtividade celular (P X ) de 185 mg L -1 d -1 , velocidade específica máxima de crescimento (µ m ) de 0,42 d -1 , fator de conversão de nitrogênio em células (Y X/N ) de 8,85 mg mg -1 , teor final de clorofila (CL f ) de 4,3 mg g -1 , e teores de proteínas (PTN) e lipídeos (LIP) de 35 e 21 %, respectivamente. Com a finalidade de estimular o crescimento celular de A. platensis, optou-se por aumentar o valor da intensidade luminosa de 156 para 192 ou 252 ± 20 µmol fótons m -2 s -1 no 5º, 8º ou 11º dia de cultivo. Os melhores resultados cinéticos (X m = 3954 mg L -1 , P X = 253 mg L -1 d -1 ) e de conteúdo da biomassa (CL f = 4,2 mg g -1 , PTN = 28 %, LIP = 19 %) foram obtidos com aumento da intensidade luminosa para 192 ± 20 µmol fótons m -2 s -1 no 8º dia de cultivo. Os ensaios realizados sob tais condições otimizadas, porém com dióxido de carbono proveniente da fermentação alcoólica, levaram à obtenção dos seguintes resultados: X m = 3298 mg L -1 , P X = 206 mg L -1 d -1 , CL f = 4,0 mg g -1 , PTN = 28 %, LIP = 17 %. Por fim, conclui-se viável a utilização do gás carbônico resultante da fermentação alcoólica no cultivo de A. platensis. Sugere-se um estudo mais aprofundado da influência desse gás efluente industrial no metabolismo e crescimento de tal microrganismo fotossintetizante, a fim de possibilitar a obtenção de resultados de concentração e produtividade celulares tão altos quanto aqueles obtidos quando do uso de gás carbônico puro. The main objective of this work was the evaluation of the potential of using carbon dioxide from alcoholic fermentation on Spirulina (Arthrospira) platensis cultivation, aiming to prove the feasibility of applying an effluent gas in the production of high added-value microbial biomass. In order to do so, the cyanobacterium was cultivated in laboratorial-scale open raceway tanks at temperature 30 ± 1 °C and light intensity 156 ± 20 µmol photons ...

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Control of fed-batch fermentations

Cited by 251 publications

References 49 publications

High Glycolytic Flux Improves Pyruvate Production by a Metabolically Engineered Escherichia coli Strain

High Glycolytic Flux Improves Pyruvate Production by a Metabolically Engineered Escherichia coli Strain

High purity recombinant human growth hormone (rhGH) expression in Escherichia coli under phoA promoter

Avaliação da utilização do dióxido de carbono proveniente da fermentação alcoólica no cultivo de Spirulina (Arthrospira) platensis: uso simultâneo de nitrato de sódio e sulfato de amônio como fontes de nitrogênio em fotobiorreator aberto

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