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Environmental Research Jul 2023Ruminant animals house a dense and diverse community of microorganisms in their rumen, an enlarged compartment in their stomach, which provides a supportive environment... (Review)
Review
Ruminant animals house a dense and diverse community of microorganisms in their rumen, an enlarged compartment in their stomach, which provides a supportive environment for the storage and microbial fermentation of ingested feeds dominated by plant materials. The rumen microbiota has acquired diverse and functionally overlapped enzymes for the degradation of plant cell wall polysaccharides. In rumen Bacteroidetes, enzymes involved in degradation are clustered into polysaccharide utilization loci to facilitate coordinated expression when target polysaccharides are available. Firmicutes use free enzymes and cellulosomes to degrade the polysaccharides. Fibrobacters either aggregate lignocellulose-degrading enzymes on their cell surface or release them into the extracellular medium in membrane vesicles, a mechanism that has proven extremely effective in the breakdown of recalcitrant cellulose. Based on current metagenomic analyses, rumen Bacteroidetes and Firmicutes are categorized as generalist microbes that can degrade a wide range of polysaccharides, while other members adapted toward specific polysaccharides. Particularly, there is ample evidence that Verrucomicrobia and Spirochaetes have evolved enzyme systems for the breakdown of complex polysaccharides such as xyloglucans, peptidoglycans, and pectin. It is concluded that diversity in degradation mechanisms is required to ensure that every component in feeds is efficiently degraded, which is key to harvesting maximum energy by host animals.
Topics: Animals; Metagenome; Rumen; Lignin; Bacteria; Polysaccharides; Bacteroidetes
PubMed: 37086884
DOI: 10.1016/j.envres.2023.115925 -
Microorganisms Jun 2023Biotechnology has a high potential to substantially contribute to a low-carbon society. Several green processes are already well established, utilizing the unique... (Review)
Review
Biotechnology has a high potential to substantially contribute to a low-carbon society. Several green processes are already well established, utilizing the unique capacity of living cells or their instruments. Beyond that, the authors believe that there are new biotechnological procedures in the pipeline which have the momentum to add to this ongoing change in our economy. Eight promising biotechnology tools were selected by the authors as potentially impactful game changers: (i) the Wood-Ljungdahl pathway, (ii) carbonic anhydrase, (iii) cutinase, (iv) methanogens, (v) electro-microbiology, (vi) hydrogenase, (vii) cellulosome and, (viii) nitrogenase. Some of them are fairly new and are explored predominantly in science labs. Others have been around for decades, however, with new scientific groundwork that may rigorously expand their roles. In the current paper, the authors summarize the latest state of research on these eight selected tools and the status of their practical implementation. We bring forward our arguments on why we consider these processes real game changers.
PubMed: 37375016
DOI: 10.3390/microorganisms11061514 -
Science (New York, N.Y.) Mar 2024Humans, like all mammals, depend on the gut microbiome for digestion of cellulose, the main component of plant fiber. However, evidence for cellulose fermentation in the...
Humans, like all mammals, depend on the gut microbiome for digestion of cellulose, the main component of plant fiber. However, evidence for cellulose fermentation in the human gut is scarce. We have identified ruminococcal species in the gut microbiota of human populations that assemble functional multienzymatic cellulosome structures capable of degrading plant cell wall polysaccharides. One of these species, which is strongly associated with humans, likely originated in the ruminant gut and was subsequently transferred to the human gut, potentially during domestication where it underwent diversification and diet-related adaptation through the acquisition of genes from other gut microbes. Collectively, these species are abundant and widespread among ancient humans, hunter-gatherers, and rural populations but are rare in populations from industrialized societies thus indicating potential disappearance in response to the westernized lifestyle.
Topics: Humans; Cellulose; Gastrointestinal Microbiome; Ruminococcus; Dietary Fiber; Phylogeny; Industrial Development
PubMed: 38484069
DOI: 10.1126/science.adj9223 -
Nature Communications Oct 2023Bacterial σ factors of the σ-family are widespread in Bacilli and Clostridia and are involved in the heat shock response, iron metabolism, virulence, and carbohydrate...
Bacterial σ factors of the σ-family are widespread in Bacilli and Clostridia and are involved in the heat shock response, iron metabolism, virulence, and carbohydrate sensing. A multiplicity of σ paralogues in some cellulolytic bacteria have been shown to be responsible for the regulation of the cellulosome, a multienzyme complex that mediates efficient cellulose degradation. Here, we report two structures at 3.0 Å and 3.3 Å of two transcription open complexes formed by two σ factors, SigI1 and SigI6, respectively, from the thermophilic, cellulolytic bacterium, Clostridium thermocellum. These structures reveal a unique, hitherto-unknown recognition mode of bacterial transcriptional promoters, both with respect to domain organization and binding to promoter DNA. The key characteristics that determine the specificities of the σ paralogues were further revealed by comparison of the two structures. Consequently, the σ factors represent a distinct set of the σ-family σ factors, thus highlighting the diversity of bacterial transcription.
Topics: Bacterial Proteins; Clostridium; Clostridium thermocellum; Promoter Regions, Genetic; Bacteria; Sigma Factor
PubMed: 37833284
DOI: 10.1038/s41467-023-41796-4 -
Scientific Data May 2024The genus Clostridium is a large and diverse group within the Bacillota (formerly Firmicutes), whose members can encode useful complex traits such as solvent production,...
The genus Clostridium is a large and diverse group within the Bacillota (formerly Firmicutes), whose members can encode useful complex traits such as solvent production, gas-fermentation, and lignocellulose breakdown. We describe 270 genome sequences of solventogenic clostridia from a comprehensive industrial strain collection assembled by Professor David Jones that includes 194 C. beijerinckii, 57 C. saccharobutylicum, 4 C. saccharoperbutylacetonicum, 5 C. butyricum, 7 C. acetobutylicum, and 3 C. tetanomorphum genomes. We report methods, analyses and characterization for phylogeny, key attributes, core biosynthetic genes, secondary metabolites, plasmids, prophage/CRISPR diversity, cellulosomes and quorum sensing for the 6 species. The expanded genomic data described here will facilitate engineering of solvent-producing clostridia as well as non-model microorganisms with innately desirable traits. Sequences could be applied in conventional platform biocatalysts such as yeast or Escherichia coli for enhanced chemical production. Recently, gene sequences from this collection were used to engineer Clostridium autoethanogenum, a gas-fermenting autotrophic acetogen, for continuous acetone or isopropanol production, as well as butanol, butanoic acid, hexanol and hexanoic acid production.
Topics: Clostridium; Phylogeny; Genome, Bacterial; Solvents; Fermentation
PubMed: 38693191
DOI: 10.1038/s41597-024-03210-6 -
Protein Expression and Purification Oct 2023Anaerobic fungi (Neocallimastigomycetes) found in the guts of herbivores are biomass deconstruction specialists with a remarkable ability to extract sugars from...
Anaerobic fungi (Neocallimastigomycetes) found in the guts of herbivores are biomass deconstruction specialists with a remarkable ability to extract sugars from recalcitrant plant material. Anaerobic fungi, as well as many species of anaerobic bacteria, deploy multi-enzyme complexes called cellulosomes, which modularly tether together hydrolytic enzymes, to accelerate biomass hydrolysis. While the majority of genomically encoded cellulosomal genes in Neocallimastigomycetes are biomass degrading enzymes, the second largest family of cellulosomal genes encode spore coat CotH domains, whose contribution to fungal cellulosome and/or cellular function is unknown. Structural bioinformatics of CotH proteins from the anaerobic fungus Piromyces finnis shows anaerobic fungal CotH domains conserve key ATP and Mg binding motifs from bacterial Bacillus CotH proteins known to act as protein kinases. Experimental characterization further demonstrates ATP hydrolysis activity in the presence and absence of substrate from two cellulosomal P. finnis CotH proteins when recombinantly produced in E. coli. These results present foundational evidence for CotH activity in anaerobic fungi and provide a path towards elucidating the functional contribution of this protein family to fungal cellulosome assembly and activity.
Topics: Cellulosomes; Escherichia coli; Anaerobiosis; Bacterial Proteins; Spores; Adenosine Triphosphate; Fungi
PubMed: 37331410
DOI: 10.1016/j.pep.2023.106323 -
Heliyon Jan 2024Cellulose degradation is a critical process in soil ecosystems, playing a vital role in nutrient cycling and organic matter decomposition. Enzymatic degradation of...
Cellulose degradation is a critical process in soil ecosystems, playing a vital role in nutrient cycling and organic matter decomposition. Enzymatic degradation of cellulosic biomass is the most sustainable and green method of producing liquid biofuel. It has gained intensive research interest with future perspective as the majority of terrestrial lignocellulose biomass has a great potential to be used as a source of bioenergy. However, the recalcitrant nature of lignocellulose limits its use as a source of energy. Noteworthy enough, enzymatic conversion of cellulose biomass could be a leading future technology. Fungal enzymes play a central role in cellulose degradation. Our understanding of fungal cellulases has substantially redirected in the past few years with the discovery of a new class of enzymes and Cellulosome. Efforts have been made from time to time to develop an economically viable method of cellulose degradation. This review provides insights into the current state of knowledge regarding cellulose degradation in soil and identifies areas where further research is needed.
PubMed: 38234915
DOI: 10.1016/j.heliyon.2024.e24022 -
Molecules (Basel, Switzerland) May 2024The hydrolysis and biotransformation of lignocellulose, i.e., biorefinery, can provide human beings with biofuels, bio-based chemicals, and materials, and is an... (Review)
Review
The hydrolysis and biotransformation of lignocellulose, i.e., biorefinery, can provide human beings with biofuels, bio-based chemicals, and materials, and is an important technology to solve the fossil energy crisis and promote global sustainable development. Biorefinery involves steps such as pretreatment, saccharification, and fermentation, and researchers have developed a variety of biorefinery strategies to optimize the process and reduce process costs in recent years. Lignocellulosic hydrolysates are platforms that connect the saccharification process and downstream fermentation. The hydrolysate composition is closely related to biomass raw materials, the pretreatment process, and the choice of biorefining strategies, and provides not only nutrients but also possible inhibitors for downstream fermentation. In this review, we summarized the effects of each stage of lignocellulosic biorefinery on nutrients and possible inhibitors, analyzed the huge differences in nutrient retention and inhibitor generation among various biorefinery strategies, and emphasized that all steps in lignocellulose biorefinery need to be considered comprehensively to achieve maximum nutrient retention and optimal control of inhibitors at low cost, to provide a reference for the development of biomass energy and chemicals.
Topics: Lignin; Hydrolysis; Biomass; Fermentation; Biofuels; Nutrients
PubMed: 38792135
DOI: 10.3390/molecules29102275 -
Applied Microbiology and Biotechnology Jul 2023The development of multifunctional particles using polymeric scaffolds is an emerging technology for many nanobiotechnological applications. Here we present a system for...
The development of multifunctional particles using polymeric scaffolds is an emerging technology for many nanobiotechnological applications. Here we present a system for the production of multifunctional complexes, based on the high affinity non-covalent interaction of cohesin and dockerin modules complementary fused to decameric Brucella abortus lumazine synthase (BLS) subunits, and selected target proteins, respectively. The cohesin-BLS scaffold was solubly expressed in high yield in Escherichia coli, and revealed a high thermostability. The production of multienzymatic particles using this system was evaluated using the catalytic domain of Cellulomonas fimi endoglucanase CenA recombinantly fused to a dockerin module. Coupling of the enzyme to the scaffold was highly efficient and occurred with the expected stoichiometry. The decavalent enzymatic complexes obtained showed higher cellulolytic activity and association to the substrate compared to equivalent amounts of the free enzyme. This phenomenon was dependent on the multiplicity and proximity of the enzymes coupled to the scaffold, and was attributed to an avidity effect in the polyvalent enzyme interaction with the substrate. Our results highlight the usefulness of the scaffold presented in this work for the development of multifunctional particles, and the improvement of lignocellulose degradation among other applications. KEY POINTS: • New system for multifunctional particle production using the BLS scaffold • Higher cellulolytic activity of polyvalent endoglucanase compared to the free enzyme • Amount of enzyme associated to cellulose is higher for the polyvalent endoglucanase.
Topics: Cellulase; Cellulomonas; Catalytic Domain; Bacterial Proteins
PubMed: 37212884
DOI: 10.1007/s00253-023-12581-6 -
Frontiers in Fungal Biology 2023Anaerobic fungi produce biomass-degrading enzymes and natural products that are important to harness for several biotechnology applications. Although progress has been...
Anaerobic fungi produce biomass-degrading enzymes and natural products that are important to harness for several biotechnology applications. Although progress has been made in the development of methods for extracting nucleic acids for genomic and transcriptomic sequencing of these fungi, most studies are limited in that they do not sample multiple fungal growth phases in batch culture. In this study, we establish a method to harvest RNA from fungal monocultures and fungal-methanogen co-cultures, and also determine an optimal time frame for high-quality RNA extraction from anaerobic fungi. Based on RNA quality and quantity targets, the optimal time frame in which to harvest anaerobic fungal monocultures and fungal-methanogen co-cultures for RNA extraction was 2-5 days of growth post-inoculation. When grown on cellulose, the fungal strain cocultivated with the methanogen upregulated genes encoding fungal carbohydrate-active enzymes and other cellulosome components relative to fungal monocultures during this time frame, but expression patterns changed at 24-hour intervals throughout the fungal growth phase. These results demonstrate the importance of establishing methods to extract high-quality RNA from anaerobic fungi at multiple time points during batch cultivation.
PubMed: 37746117
DOI: 10.3389/ffunb.2023.1171100