Clostridium thermocellum: A microbial platform for high-value chemical production from lignocellulose

Mazzoli, Roberto; Olson, Daniel G.

doi:10.1016/bs.aambs.2020.07.004

Cited by 30 publications

(27 citation statements)

References 144 publications

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“…Interestingly, the products of Clocel_4197 and Clocel_1949 were not identified in the proteome of glucose- or avicel-grown C. cellulovorans which suggests that they are not biosynthesized in high amounts in physiological conditions ( Usai et al, 2020 ). A number of studies have identified ADHs as involved in conferring alcohol resistance to bacteria since they were overexpressed under alcohol-stress ( Rutherford et al, 2010 ) or their deletion or mutation in cofactor (NADH/NADPH) specificity improved alcohol tolerance ( Brown et al, 2011 ; Dai et al, 2012 ; Tian et al, 2019 ; Mazzoli and Olson, 2020 ). Recently, butanol dehydrogenase activity has been hypothesized for the product of both Clocel_4197 and Clocel_1949 ( Wen et al, 2019 ) although this requires experimental confirmation.…”

Section: Resultsmentioning

confidence: 99%

“…By using metabolic engineering, recombinant C. cellulovorans strains have been obtained which can produce about 4 g/L and 4.96 g/L of butanol through direct fermentation of crystalline cellulose ( Bao et al, 2019 ) and alkali-extracted corn cobs ( Wen et al, 2020 ), respectively. These butanol titers are the highest reported so far for direct fermentation of plant biomass using a single microorganism ( Mazzoli and Olson, 2020 ).…”

Section: Introductionmentioning

confidence: 81%

“…In fact, most metabolic reactions leading to conversion of acetyl-CoA to butyrate are in common with butanol biosynthesis. Butyrate production is widespread in solventogenic clostridia such as C. acetobutylicum and C. beijerinckii but is absent in most cellulolytic clostridia such as C. thermocellum and C. cellulolyticum ( Mazzoli and Olson, 2020 ). By using metabolic engineering, recombinant C. cellulovorans strains have been obtained which can produce about 4 g/L and 4.96 g/L of butanol through direct fermentation of crystalline cellulose ( Bao et al, 2019 ) and alkali-extracted corn cobs ( Wen et al, 2020 ), respectively.…”

Section: Introductionmentioning

confidence: 99%

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Clostridium cellulovorans Proteomic Responses to Butanol Stress

Costa

Usai

et al. 2021

Front. Microbiol.

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Combination of butanol-hyperproducing and hypertolerant phenotypes is essential for developing microbial strains suitable for industrial production of bio-butanol, one of the most promising liquid biofuels. Clostridium cellulovorans is among the microbial strains with the highest potential for direct production of n-butanol from lignocellulosic wastes, a process that would significantly reduce the cost of bio-butanol. However, butanol exhibits higher toxicity compared to ethanol and C. cellulovorans tolerance to this solvent is low. In the present investigation, comparative gel-free proteomics was used to study the response of C. cellulovorans to butanol challenge and understand the tolerance mechanisms activated in this condition. Sequential Window Acquisition of all Theoretical fragment ion spectra Mass Spectrometry (SWATH-MS) analysis allowed identification and quantification of differentially expressed soluble proteins. The study data are available via ProteomeXchange with the identifier PXD024183. The most important response concerned modulation of protein biosynthesis, folding and degradation. Coherent with previous studies on other bacteria, several heat shock proteins (HSPs), involved in protein quality control, were up-regulated such as the chaperones GroES (Cpn10), Hsp90, and DnaJ. Globally, our data indicate that protein biosynthesis is reduced, likely not to overload HSPs. Several additional metabolic adaptations were triggered by butanol exposure such as the up-regulation of V- and F-type ATPases (involved in ATP synthesis/generation of proton motive force), enzymes involved in amino acid (e.g., arginine, lysine, methionine, and branched chain amino acids) biosynthesis and proteins involved in cell envelope re-arrangement (e.g., the products of Clocel_4136, Clocel_4137, Clocel_4144, Clocel_4162 and Clocel_4352, involved in the biosynthesis of saturated fatty acids) and a redistribution of carbon flux through fermentative pathways (acetate and formate yields were increased and decreased, respectively). Based on these experimental findings, several potential gene targets for metabolic engineering strategies aimed at improving butanol tolerance in C. cellulovorans are suggested. This includes overexpression of HSPs (e.g., GroES, Hsp90, DnaJ, ClpC), RNA chaperone Hfq, V- and F-type ATPases and a number of genes whose function in C. cellulovorans is currently unknown.

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Section: Resultsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 81%

Section: Introductionmentioning

confidence: 99%

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Clostridium cellulovorans Proteomic Responses to Butanol Stress

Costa

Usai

et al. 2021

Front. Microbiol.

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“…In addition to mesophilic Clostridia, the thermophilic Clostridia are also promising hosts for bio-ester production, because higher temperature could facilitate the downstream ester separation ( Mazzoli and Olson, 2020 ). Seo et al (2019) demonstrated that a thermostable chloramphenicol acetyltransferase from Staphylococcus aureus (CATSa) can work as a potential AAT in Clostridium thermocellum .…”

Section: Alcohol Acyltransferase-mediated Ester Synthesis In Clostridiamentioning

confidence: 99%

Developing Clostridia as Cell Factories for Short- and Medium-Chain Ester Production

Wang

Makishah

et al. 2021

Front. Bioeng. Biotechnol.

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Short- and medium-chain volatile esters with flavors and fruity fragrances, such as ethyl acetate, butyl acetate, and butyl butyrate, are usually value-added in brewing, food, and pharmacy. The esters can be naturally produced by some microorganisms. As ester-forming reactions are increasingly deeply understood, it is possible to produce esters in non-natural but more potential hosts. Clostridia are a group of important industrial microorganisms since they can produce a variety of volatile organic acids and alcohols with high titers, especially butanol and butyric acid through the CoA-dependent carbon chain elongation pathway. This implies sufficient supplies of acyl-CoA, organic acids, and alcohols in cells, which are precursors for ester production. Besides, some Clostridia could utilize lignocellulosic biomass, industrial off-gas, or crude glycerol to produce other branched or straight-chain alcohols and acids. Therefore, Clostridia offer great potential to be engineered to produce short- and medium-chain volatile esters. In the review, the efforts to produce esters from Clostridia via in vitro lipase-mediated catalysis and in vivo alcohol acyltransferase (AAT)-mediated reaction are comprehensively revisited. Besides, the advantageous characteristics of several Clostridia and clostridial consortia for bio-ester production and the driving force of synthetic biology to clostridial chassis development are also discussed. It is believed that synthetic biotechnology should enable the future development of more effective Clostridia for ester production.

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“…C. thermocellum can hydrolyze all three types of cell wall polymers, cellulose, hemicellulose (e.g., xylan) and pectin; the latter being a major component of the "glue" that holds the cells of plant biomass together [8][9][10]. Limitations in the use of C. thermocellum for the production of biofuels include the inability of native strains to utilize C5 sugars and a relatively low tolerance to ethanol [11].…”

Section: Introductionmentioning

confidence: 99%

Composition and yield of non-cellulosic and cellulosic sugars in soluble and particulate fractions during consolidated bioprocessing of poplar biomass by Clostridium thermocellum

Biswal¹,

Hengge

Black³

et al. 2021

Preprint

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Background Terrestrial plant biomass is the primary renewable carbon feedstock for enabling transition to a sustainable bioeconomy. Consolidated bioprocessing (CBP) by the cellulolytic thermophile Clostridium thermocellum offers a single step microbial platform for production of biofuels and biochemicals via simultaneous solubilization of carbohydrates from lignocellulosic biomass and conversion to products. Here, solubilization of cell wall cellulosic, hemicellulosic, and pectic polysaccharides in the liquor and solid residues generated during CBP of poplar biomass by C. thermocellum was analyzed. Results The total amount of biomass solubilized in the C. thermocellum fermentation platform was 5.8, 10.3, and 13.7% of milled non-pretreated poplar after 24, 48, and 120 h, respectively. When combined, these results demonstrate solubilization of 24% cellulose and 17% non-cellulosic sugars after 120 h, consistent with prior reports. The solubilization of non-cellulosic sugars by C. thermocellum compared to uninoculated control fermentations was 13 to 36% of arabinose (Ara), xylose (Xyl), galactose (Gal), mannose (Man), and glucose (Glc); and 15% and 3% of fucose and glucuronic acid, respectively. No rhamnose was solubilized and 71% of the galacturonic acid (GalA) was solubilized. These results indicate that C. thermocellum may be selective for the types and/or rate of solubilization of the non-cellulosic wall polymers. Xyl, Man, and Glc were found to accumulate in the fermentation liquor at levels greater than in uninoculated control fermentations, whereas Ara and Gal did not accumulate, suggesting that C. thermocellum solubilizes both hemicelluloses and pectins but utilizes them differently. After five days of fermentation, the amount of Rha in the solid residues increased 27%, indicating that the Rha-containing polymers (i.e., rhamnogalacturonan I, RG-I) were not effectively solubilized by C. thermocellum CBP. Comparison of the sugars in the liquor versus solid residue showed that C. thermocellum solubilized hemicellulosic xylan and mannan but did not fully utilize them, solubilized and appeared to utilize pectic homogalacturonan, and did not solubilize pectic RG-I. Conclusions The significant increase in Rha in poplar solid residues following CBP indicates that C. thermocellum did not solubilize RG-I. These results support the hypothesis that this pectic glycan may be a barrier for efficient solubilization of poplar by C. thermocellum.

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Clostridium thermocellum: A microbial platform for high-value chemical production from lignocellulose

Cited by 30 publications

References 144 publications

Clostridium cellulovorans Proteomic Responses to Butanol Stress

Clostridium cellulovorans Proteomic Responses to Butanol Stress

Developing Clostridia as Cell Factories for Short- and Medium-Chain Ester Production

Composition and yield of non-cellulosic and cellulosic sugars in soluble and particulate fractions during consolidated bioprocessing of poplar biomass by Clostridium thermocellum

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