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L-ornithine, a valuable non-protein amino acid, has a wide range of applications in the pharmaceutical and food industries. Currently, microbial fermentation is a promising, sustainable, and environment-friendly method to produce L-ornithine. However, the industrial production capacity of L-ornithine by microbial fermentation is low and rarely meet...
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Context 1
... the production titer of L-ornithine was low when S. cerevisiae was used as a chassis microorganism and hence, the process could not be scaled up to industrial production. As shown in Table 1, C. glutamicum is predominantly used to produce L-ornithine. C. glutamicum, a gram positive bacterium, was intensively engineered by mutation breeding to enable utilization of a broad spectrum of carbon sources to produce desired chemical compounds (Jeandet et al., 2018;Becker and Wittmann, 2019;Kim et al., 2019). ...Context 2
... the production titer of L-ornithine was low when S. cerevisiae was used as a chassis microorganism and hence, the process could not be scaled up to industrial production. As shown in Table 1, C. glutamicum is predominantly used to produce L-ornithine. C. glutamicum, a gram positive bacterium, was intensively engineered by mutation breeding to enable utilization of a broad spectrum of carbon sources to produce desired chemical compounds (Jeandet et al., 2018;Becker and Wittmann, 2019;Kim et al., 2019). ...Similar publications
Diamines are important monomers for polyamide plastics, including 1,3-diaminopropane, 1,4-diaminobutane, 1,5-diaminopentane, 1,6-diaminohexane, etc. With increasing attention on environmental problems and green sustainable development, utilizing renewable raw materials for the synthesis of diamines is crucial for the establishment of a sustainable...
Citations
... Corynebacterium glutamicum, a generally recognized as safe (GRAS) strain, manufactures high-valueadded compounds [28,29]. Recently, significant efforts have been directed toward engineering C. glutamicum to produce aromatic compounds [30]. ...
Background
β-Arbutin, a hydroquinone glucoside found in pears, bearberry leaves, and various plants, exhibits antioxidant, anti-inflammatory, antimicrobial, and anticancer effects. β-Arbutin has wide applications in the pharmaceutical and cosmetic industries. However, the limited availability of high-performance strains limits the biobased production of β-arbutin.
Results
This study established the β-arbutin biosynthetic pathway in C. glutamicum ATCC13032 by introducing codon-optimized ubiC, MNX1, and AS. Additionally, the production titer of β-arbutin was increased by further inactivation of csm and trpE to impede the competitive metabolic pathway. Further modification of the upstream metabolic pathway and supplementation of UDP-glucose resulted in the final engineered strain, C. glutamicum AR11, which achieved a β-arbutin production titer of 7.94 g/L in the optimized fermentation medium.
Conclusions
This study represents the first successful instance of de novo β-arbutin production in C. glutamicum, offering a chassis cell for β-arbutin biosynthesis.
... The KEGG analysis demonstrated that the symbiotic meal improved pathways related to energy metabolism, amino sugar metabolism [54], essential amino acid production and metabolism [55], degradation of sucrose [56], and the biosynthesis of ribonucleotides [57]. The predictive effects observed in amino acid and amino sugar pathways are related to improved immune function, oxidative stress, and immune response [11]. ...
Background: Chronic kidney disease increases uremic toxins concentrations, which have been associated with intestinal dysbiosis. Sorghum bicolor L. Moench has dietary fiber and bioactive compounds, while Bifidobacterium longum can promote beneficial health effects. Methods: It is a controlled, randomized, and single-blind clinical trial. Thirty-nine subjects were randomly separated into two groups: symbiotic group (SG), which received 100 mL of unfermented probiotic milk with Bifidobacterium longum strain and 40 g of extruded sorghum flakes; and the control group (CG), which received 100 mL of pasteurized milk and 40 g of extruded corn flakes for seven weeks. Results: The uremic toxins decreased, and gastrointestinal symptoms improved intragroup in the SG group. The acetic, propionic, and butyric acid production increased intragroup in the SG group. Regarding α-diversity, the Chao1 index was enhanced in the SG intragroup. The KEGG analysis revealed that symbiotic meal increased the intragroup energy and amino sugar metabolism, in addition to enabling essential amino acid production and metabolism, sucrose degradation, and the biosynthesis of ribonucleotide metabolic pathways. Conclusions: The consumption of symbiotic meal reduced BMI, improved short-chain fatty acid (SCFA) synthesis and gastrointestinal symptoms, increased diversity according to the Chao1 index, and reduced uremic toxins in chronic kidney disease patients.
... Microbial production of 2-OG from renewable resources has been explored in several bacteria and yeasts including Pseudomonas fluorescens, Serratia marcescens, Bacillus megaterium, Torulopsis glabrata, and Yarrowia lipolytica since the 1940s (Otto et al. 2011). Additionally, because 2-OG is not only the starting material for the biosynthesis of glutamate-family amino acids (proline, glutamine, and arginine) but can also be biologically converted to several industrially valuable compounds, many strain development studies to produce compounds derived from 2-OG, such as glutamate (Kimura 2003), ornithine, putrescine, citrulline (Jensen et al. 2015;Wu et al. 2019), γ-aminobutyrate (known as GABA) (Dhakal et al. 2012), and 1,4-butanediol (Cheng et al. 2021), have been reported. However, with regard to the compounds biosynthesized from 2-OG, but not via glutamate (defined as 2-OG derivatives to distinguish between that and the compounds biosynthesized via glutamate), a number of production systems targeting those are limited; only 1,4-butanediol is included in this group among the compounds enumerated above. ...
2-Oxoglutarate (2-OG) is a tricarboxylate cycle intermediate that can be biologically converted into several industrially important compounds. However, studies on the fermentative production of compounds synthesized from 2-OG, but not via glutamate (defined as 2-OG derivatives), have been limited. Herein, a system that can efficiently produce 2-hydroxyglutarate (2-HG), a 2-OG derivative biosynthesized by the hgdH-encoded NADH-dependent 2-HG dehydrogenase of Acidaminococcus fermentans, was developed as a model using Corynebacterium glutamicum. First, the D3 strain, which lacked the two NADH-consuming enzymes, lactate dehydrogenase and malate dehydrogenase, as well as isocitrate lyase, was constructed as a starting strain. Next, the growth conditions that induced the accumulation of 2-OG were investigated, and it was found that the biotin- and nitrogen-limited (B/N-limited) aerobic growth conditions were suitable for this purpose. Finally, the hgdH gene of A. fermentans became overexpressed in the D3 strain by inserting it into the intergenic regions with the strong constitutive promoter of the tuf gene of C. glutamicum; the engineered strain was cultured under the B/N-limited aerobic growth conditions. The engineered strain produced 80.1 mM 2-HG with a yield of 0.390 mol/mol glucose, which are the highest titer and yield reported thus far, to the best of our knowledge. Furthermore, reverse genetics showed that the produced 2-HG was partially exported via the YggB protein (NCgl1221 protein, a mechanosensitive channel) known as an exporter for glutamate under the conditions used herein.
Key points
• An efficient 2-HG production system was developed with Corynebacterium glutamicum.
• Biotin- and nitrogen-limited aerobic growth conditions induced 2-OG production.
• Produced 2-HG was partially excreted via the glutamate exporter, YggB.
... This would substitute the commonly used one, i.e. monosodium glutamate (MSG) since many reports confirmed its adverse effects especially in excessive amounts [23,24]. Such a natural flavor is synthesized from arginine via arginase enzyme through metabolically engineered bacterial fermentation [25]. These facts prioritize the engineering of arginase enzyme preliminary through bioinformatics tools to be used for the mentioned applications and this is the goal of this study. ...
Human arginase is a multifaceted enzyme that can be utilized for various medical and industrial applications. However, no report explored the in-silico engineering of this novel enzyme. The crystal structure of human arginase 1 was downloaded from PDB and had its quality checked prior to further analysis. Afterwards, CUPSAT and DeepDDG webservers were employed to nominate the most stable variants which were prepared by UCSF Chimera v1.16 and refined by GalaxyRefine tool. Docking (to reference substrate and inhibitor), stability confirmation and dynamics simulations were conducted. G119L was the best in all the mentioned aspects which was afterwards in silico cloned as a final step. Accordingly, G119L is a valuable arginase mutant that deserves its experimental validation.
... From the standpoint of environmentally friendly biotechnology and sustainable developments, fermentation approaches have been widely used for the fermentationbased synthesis of amino acids. [16][17][18]. ...
This study aims to use some strains (Corynebacterium glutamicum, Lactobacillus bulgaricus, and Brevibacterium linens for the production of some amino acids (lysine, methionine, threonine, arginine, tyrosine, aspartic acid, glutamic acid, alanine, and proline) on synthetic fermentation media under different conditions (incubation periods; incubation temperatures; pH values; media volume and different sources of carbon and nitrogen) on the course of fermentation reactions of amino acids, and then quantitative analysis of amino acids by HPLC for choose the best conditions for increasing production of amino acids. The results showed that in the best conditions for three strains, the highest production of amino acids was recorded for Brevibacterium linens the pH (7.5), incubation temperature (30°C), medium volume (50 ml), and incubation period (48 hr), while for Corynebacterium glutamicum observed on pH (7), incubation temperature (30°C), medium volume (50 ml) and incubation period (48 hr), however, for Lactobacillus bulgaricus observed on pH 6.5, incubation temperature 37°C, medium volume (50 ml) and incubation period (48 hr). The best carbon and nitrogen sources were recorded for glucose and ammonium sulfate for three tested strains.
... seen that L-ornithine is the key precursor for biosynthesis of L-arginine, siderophore (fusarinine C) and L-Piz. It has been reported that L-ornithine is mainly produced in two pathways: the ornithine-urea cycle (OUC) and the acetyl-glutamate cycle (AGC) [14][15][16] (Fig. 1). N-Acetyl glutamate kinase (ArgB) in AGC is one of the rate-limiting enzymes, and its activity is subject to feedback inhibition by L-Arg from OUC ( Fig. 1) [10]. ...
... Bioconversion of the agricultural by-product can provide valuable insight into its potential utilisation in various industries, such as amino acids production. Amino acids have been receiving great attention due to the increasing, multi-billion USD market demand in various industries, such as the food and feed industries [15,16]. Compared to other bio-based products which have applications as specialty chemicals or food ingredients, amino acids are one of the major pillars in the field of industrial biotechnology due to their wide range of contributions to GDP and the bio-economy. ...
... Water solubility, swelling power, and water absorption of PPS were determined as described by Atukuri et al. [29], Kaur et al. [30], and Chisenga et al. [31] with slight modifications. One gram of PPS suspended in 10 mL distilled water was subjected to heating in the water bath at various temperatures (60 • C, 70 • C, 80 • C, 90 • C, 95 • C, and 100 • C) and times (5,10,15,20,25,30 min) using the one-factor-at-a-time (OFAT) approach. The suspension was swirled continuously during the process and left to cool at room temperature until it reached ambient temperature. ...
Pineapples generate large amounts of agricultural wastes during their production. To reduce environmental impacts due to poor handling of these wastes, the underutilised pineapple plant stem (PPS), which has a high starch content, can be explored for its sugar recovery. To achieve this, gelatinisation is a key process in increasing enzymes’ susceptibility. Therefore, this study aimed to enhance glucose recovery from PPS by studying the effects of gelatinisation temperature and time on its functional properties. Afterwards, the fermentable sugar obtained was used for amino acids production by Bacillus subtilis ATCC 6051. PPS has a high gelatinisation temperature (To = 111 °C; Tp = 116 °C; Tc = 161 °C) and enthalpy (ΔH = 263.58 J/g). Both temperature and time showed significant effects on its functional properties, affecting enzymatic hydrolysis. Gelatinisation temperature of 100 °C at 15 min resulted in maximum glucose recovery of 56.81 g/L (0.81 g/g hydrolysis yield) with a 3.53-fold increment over the control. Subsequently, utilisation of PPS hydrolysate in the fermentation by B. subtilis ATCC 6051 resulted in 23.53 mg/mL amino acids being produced with productivity of 0.49 g/L/h. This opens up new opportunities for the applications of PPS as well as B. subtilis ATCC 6051 in the amino acids industry.
... At the same time, the screening strength could be adjusted by changing the frequency of rare codons and the copy numbers of markers. Although this strategy cannot be directly applied to amino acids with only one tRNA, such as l -Lys, l -His, and l -Phe, it provides a promising alternative to amino acid analog-based screening strategies in breeding amino acid overproducers [84] . ...
Microbial fermentation has contributed to 80% of global amino acid production. The key to microbial fermentation is to obtain fermentation strains with high performance to produce target amino acids with a high yield. These strains are primarily derived from screening enormous mutant libraries. Therefore, a high-throughput, rapid, accurate, and universal screening strategy for amino acid overproducers has become a guarantee for obtaining optional amino acid overproducers. In recent years, the rapid development of various novel screening strategies has been witnessed. However, proper analysis and discussion of these innovative technologies are lacking. Here we systematically reviewed recent advances in screening strategies: the auxotrophic-based strategy, the biosensor-based strategy, and the latest translation-based screening strategy. The design principle, application scope, working efficiency, screening accuracy, and universality of these strategies were discussed in detail. The potential for screening nonstandard amino acid overproducers was also analyzed. Guidance for the improvement of future screening strategies is provided in this review, which could expedite the reconstruction of amino acid overproducers and help promote the fermentation industry to reduce cost, increase yield, and improve quality.
... Biotechnology for Biofuels and Bioproducts glutamicum, a Gram-positive soil bacterium, has been used to produce l-ornithine on a commercial scale and is the most competitive l-ornithine producer [2]. With the completion of C. glutamicum sequencing and resolution of metabolic pathways, persistent efforts have been made to construct l-ornithine-producing strains over the past few years [3]. These strain breeding strategies can be summarized as follows: metabolic evolution [4,5], blocking competitive metabolic pathways [6][7][8], overexpression of key genes [9,10], removal of feedback inhibition in the l-ornithine biosynthesis pathway [11,12], increasing the supplementation of precursors by modifying the glycolysis and tricarboxylic acid cycle (TCA cycle) [13], increasing NADPH availability [10,14,15], and unblocking the secretion system by overexpression of lysE [7]. ...
Background
l -Ornithine is an important medicinal intermediate that is mainly produced by microbial fermentation using glucose as the substrate. To avoid competition with human food resources, there is an urgent need to explore alternative carbon sources for l -ornithine production. In a previous study, we constructed an engineered strain, Corynebacterium glutamicum MTL13, which produces 54.56 g/L of l -ornithine from mannitol. However, compared with the titers produced using glucose as a substrate, the results are insufficient, and further improvement is required.
Results
In this study, comparative transcriptome profiling of MTL01 cultivated with glucose or mannitol was performed to identify novel targets for engineering l -ornithine-producing strains. Guided by the transcriptome profiling results, we modulated the expression of qsuR (encoding a LysR-type regulator QsuR), prpC (encoding 2-methylcitrate synthase PrpC), pdxR (encoding a MocR-type regulator PdxR), acnR (encoding a TetR-type transcriptional regulator AcnR), CGS9114_RS08985 (encoding a hypothetical protein), and CGS9114_RS09730 (encoding a TetR/AcrR family transcriptional regulator), thereby generating the engineered strain MTL25 that can produce l -ornithine at a titer of 93.6 g/L, representing a 71.6% increase as compared with the parent strain MTL13 and the highest l -ornithine titer reported so far for C. glutamicum .
Conclusions
This study provides novel indirect genetic targets for enhancing l -ornithine accumulation on mannitol and lays a solid foundation for the biosynthesis of l -ornithine from marine macroalgae, which is farmed globally as a promising alternative feedstock.
... In these cases, only the minimal L-Arg was produced in the transformant E-SPE1-S-ArgB/C cells grown in medium containing glucose so that feedback inhibition by L-Arg could be relieved and AGC could be enhanced. In contrast, the overexpression of the gene encoding an N-acetylglutamate synthetase was reported to be an efficient strategy for improving L-ornithine accumulation in engineered C. glutamicum (Wu et al., 2020). It has been reported that L-ornithine overproducers have been developed using metabolic engineering in genetically modified C. glutamicum (Shu et al., 2018), and S. cerevisiae (Qin et al., 2015) from the biosynthesis pathway. ...
... ArgA ArgB/C ArgD ArgE ArgJ OTC ODC adequate supply of precursor glutamate is another key factor for developing high L-ornithine-producing bacterial strains by overexpression of endogenous or exogenous glutamate dehydrogenase, inhibition of the glutamate secretion system, and attenuation of α-ketoglutarate dehydrogenase (Wu et al., 2020). It has also been reported that fed-batch fermentation made the best-performing strain of E. coli WL3110 produce 42.3 ± 1.0 g/L of putrescine, the yield and putrescine production were 0.256 g/g of glucose and 1.265 g/L/h, respectively (Noh et al., 2017). ...
... Although E. coli WL3110 could produce a higher putrescine titer than the transformant E-SPE1-S-ArgB/C constructed in this study. E. coli is banned for use in the fermentation, food and pharmaceutical industries because it is an important conditional pathogen and synthesizes endotoxin (Wu et al., 2020), while different strains of Aureobasidium spp. and their bioproducts, such as pullulan and polymalate, have been widely applied to various sectors of biotechnology (Wei et al., , 2022. ...
Aureobasidium melanogenum HN6.2 is a high siderophore-producing yeast-like fungal strain. After blocking siderophore biosynthesis and attenuating the expression of the ornithine carbamoyltransferase gene (the OTC gene), the obtained D-LCFAO-cre strain produced 2.1 ± 0.02 mg of intracellular L-ornithine per mg of the protein. The overexpression of the L-ornithine decarboxylase gene (the SPE1-S gene) from Saccharomyces cerevisiae in the mutant D-LCFAO-cre could make the transformant E-SPE1-S synthesize 3.6 ± 0.1 of intracellular ornithine per mg of protein and produce 10.5 g/L of putrescine. The further overexpression of the ArgB/C gene encoding bifunctional acetylglutamate kinase/N-acetyl-gamma-glutamyl-phosphate reductase in the transformant E-SPE1-S caused the transformant E-SPE1-S-ArgB/C to accumulate L-ornithine (4.2 mg/mg protein) and to produce 21.3 g/L of putrescine. During fed-batch fermentation, the transformant E-SPE1-S-ArgB/C could produce 33.4 g/L of putrescine, the yield was 0.96 g/g of glucose, and the productivity was 0.28 g/L/h. The putrescine titer was much higher than that produced by most engineered strains obtained thus far.
