Effect of substrate loading on hydrogen production during anaerobic fermentation by Clostridium thermocellum 27405

Islam, Rumana; Cicek, Nazim; Sparling, Richard; Levin, David B.

doi:10.1007/s00253-006-0316-7

Cited by 81 publications

(39 citation statements)

References 26 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Our data suggest that conversion of pyruvate to acetyl-CoA occurs via both the PFL and POR mediated pathways in C. thermocellum, consistent with previous observations of end products [7,11,12]. Two ORF's putatively encoding ldh genes have been identifi ed (genes 345 and 1053).…”

Section: Discussionsupporting

confidence: 76%

“…C. thermocellum produces greater amounts of H 2 when cultured on cellulosic substrates compared with the soluble cellulodextran cellobiose, with an average yield of 1.55 mol H 2 /mol glucose equivalent [11,12]. We have observed the production of formate, ethanol, and acetate along with H 2 and CO 2 during exponential growth of the cells, with lactate being produced, as the cells entered stationary phase [7].…”

Section: Introductionmentioning

confidence: 74%

See 1 more Smart Citation

Pyruvate catabolism and hydrogen synthesis pathway genes of Clostridium thermocellum ATCC 27405

et al. 2008

Self Cite

View full text Add to dashboard Cite

Clostridium thermocellum is a gram-positive, acetogenic, thermophilic, anaerobic bacterium that degrades cellulose and carries out mixed product fermentation, catabolising cellulose to acetate, lactate, and ethanol under various growth conditions, with the concomitant release of H 2 and CO 2 . Very little is known about the factors that determine metabolic fl uxes infl uencing H 2 synthesis in anaerobic, cellulolytic bacteria like C. thermocellum. We have begun to investigate the relationships between genome content, gene expression, and end-product synthesis in C. thermocellum cultured under different conditions. Using bioinformatics tools and the complete C. thermocellum 27405 genome sequence, we identifi ed genes encoding key enzymes in pyruvate catabolism and H 2 -synthesis pathways, and have confi rmed transcription of these genes throughout growth on α-cellulose by reverse transcriptase polymerase chain reaction. Bioinformatic analyses revealed two putative lactate dehydrogenases, one pyruvate formate lyase, four pyruvate:formate lyase activating enzymes, and at least three putative pyruvate:ferredoxin oxidoreductase (POR) or POR-like enzymes. Our data suggests that hydrogen may be generated through the action of either a Ferredoxin (Fd)-dependent NiFe hydrogenase, often referred to as "Energy-converting Hydrogenases", or via NAD(P)Hdependent Fe-only hydrogenases which would permit H 2 production from NADH generated during the glyceraldehyde-3-phosphate dehydrogenase reaction. Furthermore, our fi ndings show the presence of a gene cluster putatively encoding a membrane integral NADH:Fd oxidoreductase, suggesting a possible mechanism in which electrons could be transferred between NADH and ferredoxin. The elucidation of pyruvate catabolism pathways and mechanisms of H 2 synthesis is the fi rst step in developing strategies to increase hydrogen yields from biomass. Our studies have outlined the likely pathways leading to hydrogen synthesis in C. thermocellum strain 27405, but the actual functional roles of these gene products during pyruvate catabolism and in H 2 synthesis remain to be elucidated, and will need to be confi rmed using both expression analysis and protein characterization.

show abstract

Section: Discussionsupporting

confidence: 76%

Section: Introductionmentioning

confidence: 74%

Pyruvate catabolism and hydrogen synthesis pathway genes of Clostridium thermocellum ATCC 27405

et al. 2008

Self Cite

View full text Add to dashboard Cite

show abstract

“…Unfortunately, current yields and production rates of ethanol and/or H 2 are low due to branched product pathways (3)(4)(5) which redirect carbon and electron flux away from a desired biofuel. These unwanted products include lactate, formate, and/or acetate (6)(7)(8), as well as secreted amino acids (9,10). Thus, redirecting carbon and electron flux away from these secreted products toward either ethanol or H 2 may improve the economic viability of biofuels production using C. thermocellum.…”

mentioning

confidence: 99%

Reassessment of the Transhydrogenase/Malate Shunt Pathway in Clostridium thermocellum ATCC 27405 through Kinetic Characterization of Malic Enzyme and Malate Dehydrogenase

Taillefer

Rydzak

Levin

et al. 2015

Appl Environ Microbiol

View full text Add to dashboard Cite

Clostridium thermocellum produces ethanol as one of its major end products from direct fermentation of cellulosic biomass. Therefore, it is viewed as an attractive model for the production of biofuels via consolidated bioprocessing. However, a better understanding of the metabolic pathways, along with their putative regulation, could lead to improved strategies for increasing the production of ethanol. In the absence of an annotated pyruvate kinase in the genome, alternate means of generating pyruvate have been sought. Previous proteomic and transcriptomic work detected high levels of a malate dehydrogenase and malic enzyme, which may be used as part of a malate shunt for the generation of pyruvate from phosphoenolpyruvate. The purification and characterization of the malate dehydrogenase and malic enzyme are described in order to elucidate their putative roles in malate shunt and their potential role in C. thermocellum metabolism. The malate dehydrogenase catalyzed the reduction of oxaloacetate to malate utilizing NADH or NADPH with a k cat of 45.8 s ؊1 or 14.9 s ؊1 , respectively, resulting in a 12-fold increase in catalytic efficiency when using NADH over NADPH. The malic enzyme displayed reversible malate decarboxylation activity with a k cat of 520.8 s ؊1 . The malic enzyme used NADP ؉ as a cofactor along with NH 4 ؉ and Mn 2؉ as activators. Pyrophosphate was found to be a potent inhibitor of malic enzyme activity, with a K i of 0.036 mM. We propose a putative regulatory mechanism of the malate shunt by pyrophosphate and NH 4 ؉ based on the characterization of the malate dehydrogenase and malic enzyme.C lostridium thermocellum is a Gram-positive, anaerobic thermophile capable of reaching one of the highest growth rates on crystalline cellulose (1, 2). Furthermore, given its native ability to produce ethanol and H 2 , C. thermocellum is seen as an attractive microorganism for the production of biofuels via consolidated bioprocessing of lignocellulosic biomass. Unfortunately, current yields and production rates of ethanol and/or H 2 are low due to branched product pathways (3-5) which redirect carbon and electron flux away from a desired biofuel. These unwanted products include lactate, formate, and/or acetate (6-8), as well as secreted amino acids (9, 10). Thus, redirecting carbon and electron flux away from these secreted products toward either ethanol or H 2 may improve the economic viability of biofuels production using C. thermocellum.A key pathway node involved in the interconversion of phosphoenolpyruvate (PEP) and pyruvate, which may catalyze either substrate level phosphorylation or transhydrogenation reactions between NADH to NADP ϩ , has been revisited with C. thermocellum (10, 11). The interconversion of PEP and pyruvate may be catalyzed using a number of different putative enzymes. In contrast to most clostridia and other fermentative ethanol and/or H 2 -producing organisms (7), C. thermocellum ATCC 27405 (GenBank accession number NC_009012.1) does not encode a pyruvate kinase, which catalyzes PE...

show abstract

“…Most of our current understanding of C. thermocellum physiology is based on fermentations with carbohydrate loadings of <10 g/L [7,[13][14][15][16][17][18][19][20][21][22][23][24][25][26][27][28][29][30][31]. There are, however, a few notable exceptions.…”

mentioning

confidence: 99%

“…Their study also included a co-culture of C. thermocellum with Thermoanaerobacterium thermosaccharolyticum that grew on 80 g/L solids (Solka-Floc) with three quarters of the substrate utilized and a 100 g/L cornstover co-culture with 37% of the substrate utilized [33]. Hogsett reported a set of sequential high solids fermentations with initial cellulose loadings of 26,47, and 72 g/L and a maximum cellulose utilization of 65 g/L [34].…”

mentioning

confidence: 99%

The exometabolome of Clostridium thermocellum reveals overflow metabolism at high cellulose loading

Holwerda

Thorne

Olson

et al. 2014

Biotechnol Biofuels

View full text Add to dashboard Cite

Background: Clostridium thermocellum is a model thermophilic organism for the production of biofuels from lignocellulosic substrates. The majority of publications studying the physiology of this organism use substrate concentrations of ≤10 g/L. However, industrially relevant concentrations of substrate start at 100 g/L carbohydrate, which corresponds to approximately 150 g/L solids. To gain insight into the physiology of fermentation of high substrate concentrations, we studied the growth on, and utilization of high concentrations of crystalline cellulose varying from 50 to 100 g/L by C. thermocellum. Results: Using a defined medium, batch cultures of C. thermocellum achieved 93% conversion of cellulose (Avicel) initially present at 100 g/L. The maximum rate of substrate utilization increased with increasing substrate loading. During fermentation of 100 g/L cellulose, growth ceased when about half of the substrate had been solubilized. However, fermentation continued in an uncoupled mode until substrate utilization was almost complete. In addition to commonly reported fermentation products, amino acids -predominantly L-valine and L-alanine -were secreted at concentrations up to 7.5 g/L. Uncoupled metabolism was also accompanied by products not documented previously for C. thermocellum, including isobutanol, meso-and RR/SS-2,3-butanediol and trace amounts of 3-methyl-1-butanol, 2-methyl-1-butanol and 1-propanol. We hypothesize that C. thermocellum uses overflow metabolism to balance its metabolism around the pyruvate node in glycolysis. Conclusions: C. thermocellum is able to utilize industrially relevant concentrations of cellulose, up to 93 g/L. We report here one of the highest degrees of crystalline cellulose utilization observed thus far for a pure culture of C. thermocellum, the highest maximum substrate utilization rate and the highest amount of isobutanol produced by a wild-type organism.

show abstract

Effect of substrate loading on hydrogen production during anaerobic fermentation by Clostridium thermocellum 27405

Cited by 81 publications

References 26 publications

Pyruvate catabolism and hydrogen synthesis pathway genes of Clostridium thermocellum ATCC 27405

Pyruvate catabolism and hydrogen synthesis pathway genes of Clostridium thermocellum ATCC 27405

Reassessment of the Transhydrogenase/Malate Shunt Pathway in Clostridium thermocellum ATCC 27405 through Kinetic Characterization of Malic Enzyme and Malate Dehydrogenase

The exometabolome of Clostridium thermocellum reveals overflow metabolism at high cellulose loading

Contact Info

Product

Resources

About