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1.
Insect Mol Biol ; 2024 Jul 03.
Artículo en Inglés | MEDLINE | ID: mdl-38961541

RESUMEN

Animal silk is economically important, while silk secretion is a complex and subtle mechanism regulated by many genes. We identified the poly (ADP-ribose) polymerase (PARP1) gene of the silkworm and successfully cloned its coding sequence (CDS) sequence. Using clustered regularly interspaced short palindromic repeat (CRISPR/Cas9) technology, we screened single guide RNA (sgRNA) with high knockout efficiency by cellular experiments and obtained PARP1 mutants by knocking out the PARP1 gene of the silkworm at the individual level. We found that the mutants mainly exhibited phenotypes such as smaller cocoon size and reduced cocoon shell rate than the wild type. We also detected the expression of silk protein genes in the mutant by quantitative real-time PCR (qPCR) and found that the expression of some silk protein genes was slightly down-regulated. Meanwhile, together with the results of transcriptomic analysis, we hypothesized that PARP1 may affect the synthesis of silk proteins, resulting in their failure to function properly. Our study may provide an important reference for future in-depth refinement of the molecular mechanism of silk protein expression in silk-producing animals, as well as a potential idea for future development of molecular breeding lines of silkworms to improve silk production.

2.
Ecol Lett ; 26(6): 942-954, 2023 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-37078102

RESUMEN

Release from enemies can lead to rapid evolution in invasive plants, including reduced metabolic investment in defence. Conversely, reassociation with enemies leads to renewed evolution of defence, but the potential costs of this evolution are poorly documented. We report increased resistance of the invader Ambrosia artemisiifolia after reassociation with a coevolved specialist herbivore, and that this increase corresponds with reduced abiotic stress tolerance. Herbivore resistance was higher, but drought tolerance was lower in plants from populations with a longer reassociation history, and this corresponded with changes in phenylpropanoids involved in insect resistance and abiotic stress tolerance. These changes were corroborated by shifts in the expression of underlying biosynthetic genes and plant anti-oxidants. Together, our findings suggest rapid evolution of plant traits after reassociation with coevolved enemies, resulting in genetically based shifts in investment between abiotic and biotic stress responses, providing insights into co-evolution, plant invasion and biological control.


Asunto(s)
Evolución Biológica , Herbivoria , Animales , Herbivoria/fisiología , Plantas , Insectos , Estrés Fisiológico
3.
Bioprocess Biosyst Eng ; 40(4): 549-557, 2017 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-27987090

RESUMEN

Biological synthesis of succinic acid at low pH values was favored since it not only decreased investment cost but also simplified downstream purification process. In this study, the feasibility of using glutamate decarboxylase system to improve succinic acid production of Escherichia coli AFP111, a succinate-producing candidate with mutations in pfl, ldhA, and ptsG, under acidic conditions was investigated. By overexpressing gadBC operon in AFP111, a recombinant named as BA201 (AFP111/pMD19T-gadBC) was constructed. Fermentation at pH 5.6 showed that 30 g L-1 glucose was consumed and 26.58 g L-1 succinic acid was produced by BA201, which was 1.22- and 1.32-fold higher than that by the control BA200 (AFP111/pMD19T) containing the empty vector. Analysis of intracellular enzymes activities and ATP concentrations revealed that the activities of key enzymes involved in glucose uptake and products synthesis and intracellular ATP levels were all increased after overexpression of gadBC which were benefit for cell metabolism under weak acidic conditions. To further improve the succinic acid titer by recombinant BA201 at pH 5.6, the extracellular glutamate concentration was optimized and the final succinic acid titer increased 20.4% to 32.01 g L-1. Besides, the fermentation time was prolonged by repetitive fermentation and additional 15.78 g L-1 succinic acid was produced by recovering cells into fresh medium. The results here demonstrated a potential strategy of overexpressing gadBC for increased succinic acid production of E. coli AFP111 under weak acidic conditions.


Asunto(s)
Proteínas de Escherichia coli , Escherichia coli , Ingeniería Metabólica/métodos , Mutación , Operón , Ácido Succínico/metabolismo , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/metabolismo
4.
J Ind Microbiol Biotechnol ; 41(1): 115-23, 2014 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-24127066

RESUMEN

AFP111 is a spontaneous mutant of Escherichia coli with mutations in the glucose-specific phosphotransferase system, pyruvate formate lyase system, and fermentative lactate dehydrogenase system, created to reduce byproduct formation and increase succinic acid accumulation. In AFP111, conversion of xylose to succinic acid only generates 1.67 ATP per xylose, but requires 2.67 ATP for xylose metabolism. Therefore, the ATP produced is not adequate to accomplish the conversion of xylose to succinic acid in chemically defined medium. An E. coli mutant was obtained by atmospheric and room-temperature plasmas and metabolic evolution strategies, which had the ability to use xylose and improve the capacity of cell growth. The concentration of ATP in the mutant was 1.33-fold higher than that in AFP111 during xylose fermentation. In addition, under anaerobic fermentation with almost 80 % xylose from corn stalk hydrolysate, a succinic acid concentration of 21.1 g l(-1) was obtained, with a corresponding yield of 76 %.


Asunto(s)
Escherichia coli/genética , Ingeniería Metabólica , Mutación , Ácido Succínico/metabolismo , Zea mays/metabolismo , Acetiltransferasas/genética , Escherichia coli/enzimología , Escherichia coli/metabolismo , Fermentación , Glucosa/metabolismo , Temperatura , Xilosa/metabolismo
5.
Int J Biol Macromol ; 256(Pt 1): 128316, 2024 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-38000606

RESUMEN

The silkworm is a lepidopteran domesticated from the wild silkworm, mostly valued for its efficient synthesis of silk protein. This species' ability to spin silk has supported the 5500-year-old silk industry and the globally known "Silk Road", making the transformation of mulberry leaves into silk of great concern. Therefore, research on the silk-related genes of silkworms and their regulatory mechanisms has attracted increasing attention. Previous studies have revealed that domestic silk gland cells are endoreduplication cells, and their high-copy genome and special chromatin conformation provide conditions for the high expression of silk proteins. In this study, we systematically investigate the expression pattern of eukaryotic initiation factors (eIFs) and identified the eIF6 as a eukaryotic translation initiation factor involved in the synthesis of silk proteins. We generated an eIF6 gene deletion mutant strain of silkworm using the CRISPR/Cas9 system and investigated the function of eIF6 in silk gland development and silk protein synthesis. The results showed that deletion of eIF6 inhibited the individual development of silkworm larvae, inhibited the development of silk glands, and significantly reduced the cocoon layer ratio. Therefore, we elucidated the function of eIF6 in the development of silk glands and the synthesis of silk proteins, which is important for further elucidation of the developmental process of silk glands and the mechanism underlying the ultra-high expression of silk proteins.


Asunto(s)
Bombyx , Animales , Bombyx/metabolismo , Seda/metabolismo , Proteínas de Insectos/genética , Proteínas de Insectos/metabolismo , Larva/genética , Larva/metabolismo
6.
Plant Sci ; 346: 112181, 2024 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-38969141

RESUMEN

White birch (Betula platyphylla Suk.) is an important pioneer tree which plays a critical role in maintaining ecosystem stability and forest regeneration. The growth of birch is dramatically inhibited by salt stress, especially the root inhibition. Salt Overly Sensitive 1 (SOS1) is the only extensively characterized Na+ efflux transporter in multiple plant species. The salt-hypersensitive mutant, sos1, display significant inhibition of root growth by NaCl. However, the role of SOS1 in birch responses to salt stress remains unclear. Here, we characterized a putative Na+/H+ antiporter BpSOS1 in birch and generated the loss-of-function mutants of the birch BpSOS1 by CRISPR/Cas9 approach. The bpsos1 mutant exhibit exceptional increased salt sensitivity which links to excessive Na+ accumulation in root, stem and old leaves. We observed a dramatic reduction of K+ contents in leaves of the bpsos1 mutant plants under salt stress. Furthermore, the Na+/K+ ratio of roots and leaves is significant higher in the bpsos1 mutants than the wild-type plants under salt stress. The ability of Na+ efflux in the root meristem zone is found to be impaired which might result the imbalance of Na+ and K+ in the bpsos1 mutants. Our findings indicate that the Na+/H+ exchanger BpSOS1 plays a critical role in birch salt tolerance by maintaining Na+ homeostasis and provide evidence for molecular breeding to improve salt tolerance in birch and other trees.


Asunto(s)
Betula , Tolerancia a la Sal , Intercambiadores de Sodio-Hidrógeno , Tolerancia a la Sal/genética , Betula/genética , Betula/fisiología , Betula/metabolismo , Intercambiadores de Sodio-Hidrógeno/metabolismo , Intercambiadores de Sodio-Hidrógeno/genética , Proteínas de Plantas/metabolismo , Proteínas de Plantas/genética , Raíces de Plantas/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/fisiología , Raíces de Plantas/crecimiento & desarrollo , Estrés Salino/genética , Sodio/metabolismo
7.
Bioresour Technol ; 245(Pt B): 1710-1717, 2017 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-28622981

RESUMEN

Succinic acid is a four-carbon dicarboxylic acid, which has attracted much interest due to its abroad usage as a precursor of many industrially important chemicals in the food, chemicals, and pharmaceutical industries. Facing the shortage of crude oil supply and demand of sustainable development, biological production of succinic acid from renewable resources has become a topic of worldwide interest. In recent decades, robust producing strain selection, metabolic engineering of model strains, and process optimization for succinic acid production have been developed. This review provides an overview of succinic acid producers and cultivation technology, highlight some of the successful metabolic engineering approaches.


Asunto(s)
Fermentación , Ácido Succínico , Carbono , Ácidos Dicarboxílicos , Ingeniería Metabólica
8.
J Chromatogr A ; 1493: 57-63, 2017 Apr 14.
Artículo en Inglés | MEDLINE | ID: mdl-28292517

RESUMEN

A fully automated in-tube solid phase microextraction/liquid chromatography-post column derivatization-mass spectrometry (in-tube SPME/LC-PCD-MS) method was developed for the analysis of urinary hexanal and heptanal. Online in-tube SPME enabled effective enrichment of the low level aldehydes and elimination of matrix interferences. PCD could be simply realized by mixing the LC elute and hydroxylamine hydrochloride (HAHC) solution with just a tee. The peak broadening and loss in separation efficiency associated with post column dead-volume could be ignored and even completely eliminated by employing suitable PCD configuration. What's more, HAHC is commercially available and quite cheap, and shows no contaminations to MS. The entire procedure, including the extraction of aldehydes by in-tube SPME, LC separation, post column derivatization and MS detection were integrated together and completely automated, offering competitive advantages in terms of rapidity, economy, reproducibility and simplicity. The developed protocol was then successfully performed to determine lung cancer biomarkers (hexanal, heptanal) levels in urine samples.


Asunto(s)
Aldehídos/orina , Cromatografía Liquida/métodos , Espectrometría de Masas/métodos , Microextracción en Fase Sólida/métodos , Biomarcadores de Tumor/orina , Cromatografía Liquida/instrumentación , Humanos , Neoplasias Pulmonares/orina , Espectrometría de Masas/instrumentación , Reproducibilidad de los Resultados , Microextracción en Fase Sólida/instrumentación
9.
Sheng Wu Gong Cheng Xue Bao ; 32(10): 1372-1380, 2016 Oct 25.
Artículo en Zh | MEDLINE | ID: mdl-29027447

RESUMEN

Hyper-osmotic stress is one of the key factors that decrease the efficiency of biological succinic acid production. To increase the osmotic stress tolerance of succinate-producing Escherichia coli, we studied the influence of IrrE, an exogenous global regulator, on cell osmotic stress resistance. Fermentation results showed that cell growth and succinic acid production by the recombinant increased under different Na+ concentrations. Meanwhile, the maximum dry cell mass, glucose consumption and succinic acid concentration increased 15.6%, 22% and 23%, respectively, when fermented in a 5-L bioreactor. Expressing IrrE improved cell resistance to hyper-osmotic stress. Further comparison of intracellular osmoprotectants (trehalose and glycerol) concentrations showed that trehalose and glycerol concentrations in the recombinant increased. This suggested that introduction of IrrE could enhance intracellular osmoprotectants accumulation which conferred cell with improved resistance to osmotic stress.


Asunto(s)
Proteínas Bacterianas/metabolismo , Deinococcus/genética , Escherichia coli/metabolismo , Presión Osmótica , Ácido Succínico/metabolismo , Proteínas Bacterianas/genética , Fermentación , Glicerol , Microbiología Industrial , Estrés Fisiológico , Trehalosa
10.
Appl Biochem Biotechnol ; 179(5): 880-94, 2016 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-27000060

RESUMEN

Endoinulinase production was achieved by heteroexpression of endoinulinase-encoding gene from Aspergillus ficuum which is an eukaryotic organism in Escherichia coli BL21 (DE3). Further analysis demonstrated that the native signal peptide existed in inu2 gene lowered the enzyme expression level. To realize extracellular accumulation of target protein and improve its expression level, native signal peptide was substituted with pelB, ompC, and pelB fusing with the native signal peptides; then, the effects on endoinulinase production were investigated. As a result, E. coli A606-3, with replacement of pelB as its signal peptide, showed the highest endoinulinase enzyme activity (75.22 U/mg). Also, it suggested that eukaryotic signal peptides have an inhibition on enzyme expression in prokaryotic organism. Moreover, the condition for inulooligosaccharide (IOS) production from inulin was optimized, and an IOS yield of 94.41 % was achieved under the condition of 15 % (w/v) inulin, purified endoinulinase dosage of 5 U/g inulin, 55 °C, and pH 4.6 for 24 h. The major products of hydrolysis of inulin were identified as DP3 to DP7.


Asunto(s)
Aspergillus/enzimología , Glicósido Hidrolasas/biosíntesis , Ingeniería Metabólica , Oligosacáridos/biosíntesis , Aspergillus/genética , Clonación Molecular , Escherichia coli/genética , Regulación Fúngica de la Expresión Génica , Glicósido Hidrolasas/genética , Hidrólisis , Inulina/química , Oligosacáridos/química , Oligosacáridos/genética , Señales de Clasificación de Proteína/genética
11.
Sheng Wu Gong Cheng Xue Bao ; 31(4): 534-41, 2015 Apr.
Artículo en Zh | MEDLINE | ID: mdl-26380410

RESUMEN

Sugarcane molasses containing large amounts of sucrose is an economical substrate for succinic acid production. However, Escherichia coli AFP111 cannot metabolize sucrose although it is a promising candidate for succinic acid production. To achieve sucrose utilizing ability, we cloned and expressed cscBKA genes encoding sucrose permease, fructokinase and invertase of non-PTS sucrose-utilization system from E. coli W in E. coli AFP111 to generate a recombinant strain AFP111/pMD19T-cscBKA. After 72 h of anaerobic fermentation of the recombinant in serum bottles, 20 g/L sucrose was consumed and 12 g/L succinic acid was produced. During dual-phase fermentation comprised of initial aerobic growth phase followed by anaerobic fermentation phase, the concentration of succinic acid from sucrose and sugarcane molasses was 34 g/L and 30 g/L, respectively, at 30 h of anaerobic phase in a 3 L fermentor. The results show that the introduction of non-PTS sucrose-utilization system has sucrose-metabolizing capability for cell growth and succinic acid production, and can use cheap sugarcane molasses to produce succinic acid.


Asunto(s)
Ingeniería Metabólica , Melaza , Saccharum/química , Ácido Succínico/química , Sacarosa/química , Reactores Biológicos , Escherichia coli/genética , Escherichia coli/metabolismo , Proteínas de Escherichia coli/genética , Fermentación , Proteínas de Transporte de Membrana/genética
12.
Bioresour Technol ; 153: 327-32, 2014 Feb.
Artículo en Inglés | MEDLINE | ID: mdl-24393713

RESUMEN

In this study, sucrose, a reproducible disaccharide extracted from plants, was used as the carbon source for the production of succinic acid by Actinobacillus succinogenes NJ113. During serum bottle fermentation, the succinic acid concentration reached 57.1g/L with a yield of 71.5%. Further analysis of the sucrose utilization pathways revealed that sucrose was transported and utilized via a sucrose phosphotransferase system, sucrose-6-phosphate hydrolase, and a fructose PTS. Compared to glucose utilization in single pathway, more pathways of A. succinogenes NJ113 are dependent on sucrose utilization. By changing the control strategy in a fed-batch culture to alleviate sucrose inhibition, 60.5g/L of succinic acid was accumulated with a yield of 82.9%, and the productivity increased by 35.2%, reaching 2.16g/L/h. Thus utilization of sucrose has considerable potential economics and environmental meaning.


Asunto(s)
Actinobacillus/metabolismo , Ácido Succínico/metabolismo , Sacarosa/metabolismo , Actinobacillus/efectos de los fármacos , Técnicas de Cultivo Celular por Lotes , Transporte Biológico/efectos de los fármacos , Carbono/farmacología , Fermentación/efectos de los fármacos , Fructosa/metabolismo , Redes y Vías Metabólicas/efectos de los fármacos , Fosfotransferasas/metabolismo
13.
Enzyme Microb Technol ; 56: 8-14, 2014 Mar 05.
Artículo en Inglés | MEDLINE | ID: mdl-24564896

RESUMEN

Succinate is not the dominant fermentation product from xylose in wild-type Escherichia coli K12. E. coli BA 203 is a lactate dehydrogenase (ldhA), pyruvate formate lyase (pflB), and phosphoenolpyruvate (PEP)-carboxylase (ppc) deletion strain. To increase succinate accumulation and reduce byproduct formation, engineered E. coli BA204, in which ATP-forming PEP-carboxykinase (PEPCK) is overexpressed in BA203, was constructed and produced 2.17-fold higher succinate yield. To further improve the biomass and the consumption rate of xylose, nicotinic acid phosphoribosyltransferase (NAPRTase), a rate limiting enzyme in the synthesis of NAD(H), was also overexpressed. Thus, co-expression of PEPCK and NAPRTase in recombinant E. coli BA209 was investigated. In BA209, the pck gene and the pncB gene each have a trc promoter, hence, both genes are well expressed. During a 72-h anaerobic fermentation in sealed bottles, the total concentration of NAD(H) in BA209 was 1.25-fold higher than that in BA204, and the NADH/NAD+ ratio decreased from 0.28 to 0.11. During the exclusively anaerobic fermentation in a 3-L bioreactor, BA209 consumed 17.1 g L⁻¹ xylose and produced 15.5 g L⁻¹ succinate. Furthermore, anaerobic fermentation of corn stalk hydrolysate contained 30.1 g L⁻¹ xylose, 2.1 g L⁻¹ glucose and 1.5 g L⁻¹ arabinose, it produced a final succinate concentration of 17.2 g L⁻¹ with a yield of 0.94 g g⁻¹ total sugars.


Asunto(s)
Bacillus subtilis/enzimología , Proteínas Bacterianas/biosíntesis , Escherichia coli K12/enzimología , Microbiología Industrial/métodos , Pentosiltransferasa/biosíntesis , Fosfoenolpiruvato Carboxiquinasa (ATP)/biosíntesis , Succinatos/metabolismo , Anaerobiosis , Bacillus subtilis/genética , Proteínas Bacterianas/genética , Proteínas Bacterianas/aislamiento & purificación , Reactores Biológicos , Clonación Molecular , Escherichia coli K12/genética , Proteínas de Escherichia coli/biosíntesis , Proteínas de Escherichia coli/genética , Proteínas de Escherichia coli/aislamiento & purificación , Fermentación , Genes Bacterianos , Genes Sintéticos , Hidrólisis , Pentosiltransferasa/genética , Pentosiltransferasa/aislamiento & purificación , Fosfoenolpiruvato Carboxiquinasa (ATP)/genética , Fosfoenolpiruvato Carboxiquinasa (ATP)/aislamiento & purificación , Extractos Vegetales/metabolismo , Brotes de la Planta , Succinatos/aislamiento & purificación , Residuos , Xilosa/metabolismo , Zea mays
14.
Sheng Wu Gong Cheng Xue Bao ; 29(10): 1386-97, 2013 Oct.
Artículo en Zh | MEDLINE | ID: mdl-24432654

RESUMEN

Succinic acid is one of the key intermediates in the tricarboxylic acid cycle (TCA)and has huge potentials in biopolymer, food, medicine applications. This article reviews recent research progress in the production of succinic acid by microbial fermentation, including discovery and screening of the succinic-acid-producing microbes, the progress of genetic engineering strategy and metabolic engineering technology for construction of succinic acid-producing strains, and fermentation process control and optimization. Finally, we discussed the limitation of current progress and proposed the future research needs for microbial production of succinic acid.


Asunto(s)
Actinobacillus/metabolismo , Anaerobiospirillum/metabolismo , Microbiología Industrial/métodos , Ingeniería Metabólica/métodos , Ácido Succínico/metabolismo , Actinobacillus/genética , Anaerobiospirillum/genética , Fermentación , Microbiología Industrial/tendencias
15.
Sheng Wu Gong Cheng Xue Bao ; 29(12): 1875-9, 2013 Dec.
Artículo en Zh | MEDLINE | ID: mdl-24660637

RESUMEN

During the anaerobic fermentation by Escherichia coli AFP111 for succinic acid production, the viable cell concentration and productivity were decreased with the raising of succinic acid concentration. In order to restore cellular succinic acid productivity and prolong fermentation time, we collected strains and refreshed medium for repetitive succinic acid production. However, productivity is lower than that in the anaerobic fermentation before reusing strains. To enhance the productivity, strains were aerobically cultivated for 3 h in pure water before anaerobic fermentation. The activities of key enzymes were enhanced for better performance in producing succinic acid at anaerobic stage. After three rounds of repetitive fermentations, succinic acid concentration and yield reached to 56.50 g/L and 90% respectively. The succinic acid productivity was 0.81 g/(L x h), which was 13% higher than the repetitive fermentations without aerobic activation of the strains.


Asunto(s)
Escherichia coli/genética , Escherichia coli/metabolismo , Fermentación , Ácido Succínico/metabolismo , Aerobiosis , Anaerobiosis , Medios de Cultivo , Ingeniería Genética , Glucosa/metabolismo , Microbiología Industrial
16.
Bioresour Technol ; 149: 84-91, 2013 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-24096277

RESUMEN

To enhance succinic acid formation during xylose fermentation in Escherichia coli, overexpression of ATP-forming phosphoenolpyruvate carboxykinase (PEPCK) from Bacillus subtilis 168 in an ldhA, pflB, and ppc deletion strain resulted in a significant increase in cell mass and succinic acid production. However, BA204 displays a low yield of glucose fermentation and sequential glucose-xylose utilization under regulation by the phosphotransferase system (PTS). To improve the capability of glucose fermentation and simultaneously consume sugar mixture for succinic acid production, a pflB, ldhA, ppc, and ptsG deletion strain overexpressing ATP-forming PEPCK, named E. coli BA305, was constructed. As a result, after 120 h fed-batch fermentation of sugarcane bagasse hydrolysate, the dry cell weight and succinic acid concentration in BA305 were 4.58 g L(-1) and 39.3 g L(-1), respectively.


Asunto(s)
Biomasa , Escherichia coli/metabolismo , Ingeniería Genética , Glucosa/metabolismo , Lignina/metabolismo , Ácido Succínico/metabolismo , Xilosa/metabolismo , Adenosina Trifosfato/metabolismo , Anaerobiosis , Técnicas de Cultivo Celular por Lotes , Reactores Biológicos , Celulosa/metabolismo , Escherichia coli/enzimología , Fermentación , Hidrólisis , Fosfoenolpiruvato Carboxilasa/metabolismo , Sistema de Fosfotransferasa de Azúcar del Fosfoenolpiruvato/metabolismo , Saccharum/química
17.
Bioresour Technol ; 135: 574-7, 2013 May.
Artículo en Inglés | MEDLINE | ID: mdl-23010211

RESUMEN

Efficient biosynthesis of succinate from a renewable biomass resource by engineered Escherichia coli is reported in this paper. Fermentation of sugarcane bagasse hydrolysate by engineered E. coli BA204, a pflB, ldhA, and ppc deletion strain overexpressing the ATP-forming phosphoenolpyruvate carboxykinase from Bacillus subtilis 168, produced a final succinate concentration of 15.85 g L(-1) with a high yield of 0.89 g L(-1) total sugar under anaerobic conditions. During dual-phase fermentations, initial aerobic growth facilitated subsequent anaerobic succinate production, with a final succinate concentration of 18.88 g L(-1) and a yield of 0.96 g g(-1) total sugar after 24 h of anaerobic fermentation. The high succinate yield from sugarcane bagasse hydrolysate demonstrated a great potential application of renewable biomass as a feedstock for the economical production of succinate using metabolically engineered E. coli.


Asunto(s)
Carbono/farmacología , Celulosa/farmacología , Escherichia coli/metabolismo , Ingeniería Metabólica/métodos , Saccharum/química , Ácido Succínico/metabolismo , Anaerobiosis/efectos de los fármacos , Técnicas de Cultivo Celular por Lotes , Reactores Biológicos/microbiología , Carbohidratos/farmacología , Escherichia coli/efectos de los fármacos , Fermentación/efectos de los fármacos , Hidrólisis/efectos de los fármacos
18.
Bioresour Technol ; 143: 405-12, 2013 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-23819977

RESUMEN

In this study, repetitive production of succinic acid from lignocellulose hydrolysates by enhancement of ATP supply in metabolically engineered E. coli is reported. Escherichia coli BA305, a pflB, ldhA, ppc, and ptsG deletion strain overexpressing ATP-forming phosphoenolpyruvate (PEP) carboxykinase (PEPCK), produced a final succinic acid concentration of 83 g L(-1) with a high yield of 0.87 g g(-1) total sugar in 36 h of three repetitive fermentations of sugarcane bagasse hydrolysate. Furthermore, simultaneous consumption of glucose and xylose was achieved, and the specific productivity and yield of succinic acid were almost maintained constant during the repetitive fermentations.


Asunto(s)
Adenosina Trifosfato/metabolismo , Escherichia coli/metabolismo , Ingeniería Genética , Lignina/metabolismo , Ácido Succínico/metabolismo , Escherichia coli/genética , Fermentación , Hidrólisis
19.
Sheng Wu Gong Cheng Xue Bao ; 28(9): 1059-69, 2012 Sep.
Artículo en Zh | MEDLINE | ID: mdl-23289308

RESUMEN

Escherichia coli NZN111 is a promising strain with ldhA and pflB genes inactivated for the production of succinic acid. However, with these mutations, NAD+ could not be regenerated from NADH, and an unbalanced NADH/NAD+ ratio eliminated cell growth and glucose utilization under anaerobic conditions. Nicotinic acid mononucleotide adenylyltransferase (NAMNAT), encoded by the nadD gene, catalyzes the reaction from nicotinic acid mononucleotide (NaMN) to nicotinic acid adenine dinucleotide (NaAD) during the synthetic pathway of NAD(H). Overexpression of the nadD gene could enhance the concentration of NAD(H) and maintain a suitable NADH/NAD+ ratio. In this study, we constructed a recombinant strain E. coli NZN111/pTrc99a-nadD, and overexpressed NAMNAT with 1.0 mmol/L of IPTG under anaerobic conditions in sealed bottles. Compared to E. coli NZN111, the concentrations of NAD+ and NADH in the recombinant strain increased by 3.21-fold and 1.67-fold, respectively. The total concentration of NAD(H) was increased by 2.63-fold, and the ratio of NADH/NAD+ decreased from 0.64 to 0.42. The recombinant strain restored the cell growth and glucose utilization under anaerobic conditions. After 72 h, the recombinant strain could consume 14.0 g/L of glucose to produce 6.23 g/L of succinic acid, and the concentration of succinic acid was 19-fold higher than in E. coli NZN111.


Asunto(s)
Escherichia coli/genética , Nicotinamida-Nucleótido Adenililtransferasa/genética , Ácido Succínico/metabolismo , Anaerobiosis , Escherichia coli/metabolismo , Glucosa/metabolismo , Mutación , NAD/metabolismo , Nicotinamida-Nucleótido Adenililtransferasa/metabolismo , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo
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