Improvement of butanol production in <i>Clostridium acetobutylicum</i> through enhancement of NAD(P)H availability

Qi, Feng; Thakker, Chandresh; Zhu, Fayin; Peña, Matthew I.; San, Ka‐Yiu; Bennett, George N.

doi:10.1007/s10295-018-2068-7

Cited by 27 publications

(27 citation statements)

References 25 publications

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“…The co-factor NADH plays an important role in butanol biosynthesis, and insufficient intracellular NADH supply of solventogenic Clostridium limits the production of butanol (Li et al 2015; Qi et al 2018). Therefore, increasing the supply of NADH is of great significance for butanol production.…”

Section: Discussionmentioning

confidence: 99%

“…For example, NADH was involved in 43 reactions in Escherichia coli and 65 reactions in Saccharomyces cerevisiae (Nielsen 2003). Therefore, redox balance is of great significance for maintaining normal cell growth and physiological metabolism, and NADH plays an important role in maintaining the redox balance (Qi et al 2018). In addition, NADH also could affect the products formation and metabolic flux redistribution.…”

Section: Introductionmentioning

confidence: 99%

“…In addition, the NADH and ATP levels of Clostridium beijerinckii NCIMB 8052 was significantly increased by insertional inactivation of the NADH-quinone oxidoreductase gene Cbei_4110, resulting in a 21.8% improved in butanol titer (Liu et al 2016). NAD(P)H availability was also improved by overexpressing FdNR (ferredoxin NAD(P) + oxidoreductase) in the C. acetobutylicum buk − strain, which resulted in an improvement in the production of butanol and the ratio of butanol/acetate (Qi et al 2018).…”

Section: Introductionmentioning

confidence: 99%

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The significance of aspartate on NAD(H) biosynthesis and ABE fermentation in Clostridium acetobutylicum ATCC 824

Liao

Yang

et al. 2019

AMB Expr

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The co-factor NADH plays an important role in butanol biosynthesis. In this study, we found that aspartate could effectively improve the butanol production of Clostridium acetobutylicum ATCC 824. Further study showed that aspartate could be used as the precursor of NADH de novo synthesis in C. acetobutylicum ATCC 824. When 2 g/L aspartate was added, the transcription levels of essential genes (nadA, nadB and nadC) for NADH de novo synthesis were significantly higher than that of without aspartate addition. The levels of intracellular NAD+, NADH, total NAD(H) and the ratio of NADH/NAD+ were also significantly increased, which were 63.9 ± 8.0%, 85.0 ± %, 77.7 ± 8.0% and 12.7 ± 2.9% higher than those of without aspartate addition, respectively. Furthermore, the butanol production was improved by overexpressing the NADH de novo synthesis genes, and the fermentation performance could be further enhanced by strengthening the VB1 biosynthesis and NADH de novo synthesis pathway simultaneously. As a result, the butanol titer of the engineered strain 824(thiCGE–nadC) reached 13.96 ± 0.11 g/L, 7.2 ± 0.4%, 18.1 ± 0.1%, 34.1 ± 0.1% higher than that of 824(thiCGE), 824(nadC) and the wild type strain, respectively. This study has a reference value for the NADH related researches of other microbes, and the engineering strategy used in this study provides a new idea for construction of efficient fuel-producing strains.

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Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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The significance of aspartate on NAD(H) biosynthesis and ABE fermentation in Clostridium acetobutylicum ATCC 824

Liao

Yang

et al. 2019

AMB Expr

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“…Brockarchaeota BDH's are homologues to sequences from obligately anaerobic, thermophilic bacteria that can degrade complex plant saccharides such as xylan (i.e Caldicoprobacter oshimai 24 and Hungateiclostridium thermocellum 25 ) or cellulose (Hungateiclostridium alkalicellulosi). To investigate if Brockarchaeota can oxidize or produce butanol, we searched for genes involved in production of butanol in two model organisms; Clostridium acetobutylicum 26,27 which is one of the few organisms that produces butanol as a fermentation product, and Saccharomyces cerevisiae 28 involved in butanol and isopropanol production. We found that Brockarchaeota genomes lack the key enzymes involved in the fermentation of pyruvate to butanol (butanal dehydrogenase, butyryl-CoA dehydrogenase, enoyl-…”

Section: Utilization Of C1 Compounds and Central Carbon Metabolismmentioning

confidence: 99%

Brockarchaeota, a novel archaeal lineage capable of methylotrophy

Anda¹,

Chen²,

Dombrowski³

et al. 2020

Preprint

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Single carbon (C1) compounds such as methanol, methylamines and formaldehyde are ubiquitous in nature and they are large components of the carbon cycle. In anoxic environments C1-utilizing microbes (methylotrophs) play an important role in controlling global carbon degradation. Currently described anaerobic methylotrophs are limited to methanogenic archaea, acetogenic bacteria, and sulfate-reducing bacteria. Here, we report the first archaeal lineage outside of methanogenic taxa that are capable of anaerobic methylotrophy. Phylogenetic analyses suggest these archaea form a new phylum within the TACK superphylum, which we propose be named Brockarchaeota. A survey revealed Brockarchaeota are globally distributed in geothermal springs. Metabolic inference from 15 metagenome-assembled genomes from hot springs and deep-sea sediments indicates that Brockarchaeota are strict anaerobes. They contain a variety C1 metabolisms including the methanol and trimethylamine methyltransferases system, the ribulose bisphosphate pathway coupled with the non-oxidative pentoses phosphate pathway, and reductive glycine pathway. Brockarchaeota have an incomplete methyl-branch of the Wood-Ljungdahl pathway probably used for formaldehyde oxidation, since they lack several core genes involved in methanogenesis including methyl-CoM reductases. Brockarchaeota also appear to play an important role in the breakdown of plant-derived polysaccharides, especially cellulose, starch and xylan. Their broad distribution and their capacity to use both C1 compounds and complex polysaccharides via anaerobic metabolism suggest that the Brockarchaeota occupy previously overlooked roles in carbon cycling.

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“…55 This is not surprising in light of a recent study that showed Ter is detrimental to ABE fermentation when introduced in C. acetobutylicum. 56 For comparison, the previously best reported butanol production in engineered acetogenic clostridia was ~2 mM. 15 Moreover, the Bcd-EtfA:EtfB complex is a delicate complex that is extremely oxygen sensitive 57 and has so far been inactive in E. coli lysates (in alignment with previous reports that highlighted difficulties expressing Bcd in E. coli), 52 highlighting an area for potential improvement of iPROBE (i.e., compatibility of E. coli lysates with non-model organisms).…”

Section: Figure 5 Cell-free Pathway Testing Combined With Data-drivementioning

confidence: 99%

In vitroprototyping and rapid optimization of biosynthetic enzymes for cellular design

Karim

Dudley

Juminaga

et al. 2019

Preprint

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Microbial cell factories offer an attractive approach for production of biobased products.Unfortunately, designing, building, and optimizing biosynthetic pathways remains a complex challenge, especially for industrially-relevant, non-model organisms. To address this challenge, we describe a platform for in vitro Prototyping and Rapid Optimization of Biosynthetic Enzymes (iPROBE). In iPROBE, cell lysates are enriched with biosynthetic enzymes by cell-free protein synthesis and then metabolic pathways are assembled in a mix-and-match fashion to assess pathway performance. We demonstrate iPROBE with two examples. First, we tested and ranked 54 different pathways for 3-hydroxybutyrate production, improving in vivo production in Clostridium by 20-fold to 14.63 ± 0.48 g/L and identifying a new biosynthetic route to (S)-(+)-1,3butanediol. Second, we used iPROBE and data-driven design to optimize a 6-step n-butanol pathway, increasing titers 4-fold across 205 pathways, and showed strong correlation between cell-free and cellular performance. We expect iPROBE to accelerate design-build-test cycles for industrial biotechnology.

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Improvement of butanol production in Clostridium acetobutylicum through enhancement of NAD(P)H availability

Cited by 27 publications

References 25 publications

The significance of aspartate on NAD(H) biosynthesis and ABE fermentation in Clostridium acetobutylicum ATCC 824

The significance of aspartate on NAD(H) biosynthesis and ABE fermentation in Clostridium acetobutylicum ATCC 824

Brockarchaeota, a novel archaeal lineage capable of methylotrophy

In vitroprototyping and rapid optimization of biosynthetic enzymes for cellular design

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