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Microbiology (Reading, England) Jul 2012Structures of free-living and protozoa-associated methanogen (PAM) communities from forage-fed cattle were investigated by comparative sequence analysis of 16S rRNA and...
Structures of free-living and protozoa-associated methanogen (PAM) communities from forage-fed cattle were investigated by comparative sequence analysis of 16S rRNA and methyl coenzyme M reductase (mcrA) gene clone libraries. The free-living and protozoa-associated communities were composed of the same three genera [namely Methanobrevibacter, Methanomicrobium and rumen cluster C (RCC), which is distantly related to Thermoplasma]; however, the distribution of the methanogen genera differed between the two communities. Despite previous reports of potential bias for the degenerate mcrA primer set, the 16S rRNA (n = 100 clones) and mcrA (n = 92 clones) gene libraries exhibited a similar distribution pattern for the three methanogenic genera. RCC was more abundant in the free-living community and represented 72 and 42 % of the 16S rRNA and mrcA gene sequences, respectively, versus 54 and 13 % of the 16S rRNA and mrcA gene sequences from the PAM community, respectively. The majority of RCC sequences from the free-living and protozoa-associated communities belonged to different species-level operational taxonomic units. Methanobrevibacter species were more abundant in the PAM community and represented 42 and 79 % of clones for the 16S rRNA and mrcA gene libraries, respectively, versus 9 and 27 % of 16S rRNA and mrcA gene clones from the free-living community, respectively. Methanomicrobium species were predominantly free-living. Primers for quantitative PCR were designed to target specific methanogen groups and used to assess the effect of a high-grain diet on methanogen species composition. Switching the ruminant diet from forage to high-grain resulted in reduced protozoal diversity, along with a profound overall reduction in the relative abundance of RCC and an increase in the relative abundance of free-living Methanobrevibacter spp. It was unclear whether the reduced abundance of RCC in grain-fed animals was due to the loss of symbiotic protozoa species or due to broader changes in the rumen environment that affected both RCC and protozoa. Importantly, results from this study emphasize the need to consider the different methanogen communities when developing strategies for mitigating methane emissions in ruminants.
Topics: Animals; Biota; Cattle; Cluster Analysis; DNA, Archaeal; DNA, Ribosomal; Methane; Methanobrevibacter; Methanomicrobiaceae; Molecular Sequence Data; Oxidoreductases; Phylogeny; RNA, Ribosomal, 16S; Rumen; Sequence Analysis, DNA
PubMed: 22539164
DOI: 10.1099/mic.0.057984-0 -
Archaea (Vancouver, B.C.) Nov 2010PeiW (UniProtKB Q7LYX0) and PeiP (UniProtKB Q77WJ4) are the two major pseudomurein endoisopeptidases (Pei) that are known to cleave pseudomurein cell-wall sacculi of the... (Review)
Review
PeiW (UniProtKB Q7LYX0) and PeiP (UniProtKB Q77WJ4) are the two major pseudomurein endoisopeptidases (Pei) that are known to cleave pseudomurein cell-wall sacculi of the members of the methanogenic orders Methanobacteriales and Methanopyrales. Both enzymes, originating from prophages specific for some methanogenic archaeal species, hydrolyze the ϵ(Ala)-Lys bond of the peptide linker between adjacent pseudomurein layers. Because lysozyme is not able to cleave the pseudomurein cell wall, the enzymes are used in protoplast preparation and in DNA isolation from pseudomurein cell-wall-containing methanogens. Moreover, PeiW increases the probe permeability ratio and enables fluorescence in situ hybridization (FISH) and catalyzed reporter deposition (CARD-) FISH experiments to be performed on these methanogens.
Topics: Endopeptidases; Methanobacteriaceae; Peptidoglycan
PubMed: 21113291
DOI: 10.1155/2010/480492 -
Nature Communications Jun 2022Archaea are common constituents of the gut microbiome of humans, ruminants, and termites but little is known about their diversity and abundance in other animals. Here,...
Archaea are common constituents of the gut microbiome of humans, ruminants, and termites but little is known about their diversity and abundance in other animals. Here, we analyse sequencing and quantification data of archaeal and bacterial 16S rRNA genes from 250 species of animals covering a large taxonomic spectrum. We detect the presence of archaea in 175 animal species belonging to invertebrates, fish, amphibians, birds, reptiles and mammals. We identify five dominant gut lineages, corresponding to Methanobrevibacter, Methanosphaera, Methanocorpusculum, Methanimicrococcus and "Ca. Methanomethylophilaceae". Some archaeal clades, notably within Methanobrevibacter, are associated to certain hosts, suggesting specific adaptations. The non-methanogenic lineage Nitrososphaeraceae (Thaumarchaeota) is frequently present in animal samples, although at low abundance, but may have also adapted to the gut environment. Host phylogeny, diet type, fibre content, and intestinal tract physiology are major drivers of the diversity and abundance of the archaeome in mammals. The overall abundance of archaea is more influenced by these factors than that of bacteria. Methanogens reducing methyl-compounds with H can represent an important fraction of the overall methanogens in many animals. Together with CO-reducing methanogens, they are influenced by diet and composition of gut bacteria. Our results provide key elements toward our understanding of the ecology of archaea in the gut, an emerging and important field of investigation.
Topics: Animals; Archaea; Bacteria; Dietary Fiber; Euryarchaeota; Mammals; Methanobacteriaceae; Methanobrevibacter; Phylogeny; RNA, Ribosomal, 16S
PubMed: 35688919
DOI: 10.1038/s41467-022-31038-4 -
Biochimica Et Biophysica Acta.... Apr 2021In methanogenic archaea, the archetypical complex of heterodisulfide reductase (HdrABC) and hydrogenase (MvhAGD) couples the endergonic reduction of CO by H to the... (Review)
Review
In methanogenic archaea, the archetypical complex of heterodisulfide reductase (HdrABC) and hydrogenase (MvhAGD) couples the endergonic reduction of CO by H to the exergonic reduction of the CoB-S-S-CoM heterodisulfide by H via flavin-based electron bifurcation. Presently known enzymes containing HdrA(BC)-like components play key roles in methanogenesis, acetogenesis, respiratory sulfate reduction, lithotrophic reduced sulfur compound oxidation, aromatic compound degradation, fermentations, and probably many further processes. This functional diversity is achieved by a modular architecture of HdrA(BC) enzymes, where a big variety of electron input/output modules may be connected either directly or via adaptor modules to the HdrA(BC) components. Many, but not all HdrA(BC) complexes are proposed to catalyse a flavin-based electron bifurcation/confurcation. Despite the availability of HdrA(BC) crystal structures, fundamental questions of electron transfer and energy coupling processes remain. Here, we address the common properties and functional diversity of HdrA(BC) core modules integrated into electron-transfer machineries of outstanding complexity.
Topics: Archaeal Proteins; Carbon Dioxide; Dinitrocresols; Hydrogen; Methanobacteriaceae; Oxidation-Reduction; Oxidoreductases
PubMed: 33460586
DOI: 10.1016/j.bbabio.2021.148379 -
BMC Microbiology Apr 2019Methane emissions from pigs account for 10% of total methane production from livestock in China. Methane emissions not only contribute to global warming, as it has 25...
BACKGROUND
Methane emissions from pigs account for 10% of total methane production from livestock in China. Methane emissions not only contribute to global warming, as it has 25 times the global warming potential (GWP) of CO, but also represent approximately 0.1~3.3% of digestive energy loss. Methanogens also play an important role in maintaining the balance of the gut microbiome. The large intestines are the main habitat for the microbiome in pigs. Thus, to better understand the mechanism of methane production and mitigation, generic-specific and physio-ecological characteristics (including redox potential (Eh), pH and volatile fatty acids (VFAs)) and methanogens in the large intestine of pig were studied in this paper. Thirty DLY finishing pigs with the same diet and feeding conditions were selected for this experiment.
RESULT
A total of 219 clones were examined using the methyl coenzyme reductase subunit A gene (mcrA) and assigned to 43 operational taxonomic units (OTUs) based on a 97% species-level identity criterion. The family Methanobacteriaceae was the dominant methanogen in colonic digesta of finishing pigs, accounting for approximately 70.6% of the identified methanogens, and comprised mainly the genera Methanobrevibacter (57%) and Methanosphaera (14%). The order Methanomassiliicoccales, classified as an uncultured taxonomy, accounted for 15.07%. The methanogenic archaeon WGK1 and unclassified Methanomicrobiales belonging to the order of Methanomicrobiales accounted for 4.57 and 1.37%, respectively. The Eh was negative and within the range - 297.00~423.00 mV and the pH was within the range 5.04~6.97 in the large intestine. The populations of total methanogens and Methanobacteriales were stable in different parts of the large intestine according to real-time PCR.
CONCLUSION
The major methanogen in the large intestine of finishing pigs was Methanobrevibacter. The seventh order Methanomassiliicoccales and species Methanosphaera stadtmanae present in the large intestine of pigs might contribute to the transfer of hydrogen and fewer methane emissions. The redox potential (Eh) was higher in the large intestine of finishing pigs, which had a positive correlation with the population of Methanobacteriale.
Topics: Animals; China; Colon; DNA Restriction Enzymes; DNA, Ribosomal; Fatty Acids, Volatile; Female; Gastrointestinal Microbiome; Hydrogen-Ion Concentration; Intestine, Large; Male; Methane; Methanobacteriaceae; Methanobrevibacter; Oxidation-Reduction; Phylogeny; RNA, Ribosomal, 16S; Swine
PubMed: 31035941
DOI: 10.1186/s12866-019-1459-x -
The Biochemical Journal Aug 2020The DNA helicase Large helicase-related (Lhr) is present throughout archaea, including in the Asgard and Nanoarchaea, and has homologues in bacteria and eukaryotes. It...
The DNA helicase Large helicase-related (Lhr) is present throughout archaea, including in the Asgard and Nanoarchaea, and has homologues in bacteria and eukaryotes. It is thought to function in DNA repair but in a context that is not known. Our data show that archaeal Lhr preferentially targets DNA replication fork structures. In a genetic assay, expression of archaeal Lhr gave a phenotype identical to the replication-coupled DNA repair enzymes Hel308 and RecQ. Purified archaeal Lhr preferentially unwound model forked DNA substrates compared with DNA duplexes, flaps and Holliday junctions, and unwound them with directionality. Single-molecule FRET measurements showed that binding of Lhr to a DNA fork causes ATP-independent distortion and base-pair melting at, or close to, the fork branchpoint. ATP-dependent directional translocation of Lhr resulted in fork DNA unwinding through the 'parental' DNA strands. Interaction of Lhr with replication forks in vivo and in vitro suggests that it contributes to DNA repair at stalled or broken DNA replication.
Topics: Archaeal Proteins; DNA Helicases; DNA Repair; DNA Replication; DNA, Archaeal; DNA, Single-Stranded; Methanobacteriaceae; Protein Conformation
PubMed: 32706021
DOI: 10.1042/BCJ20200379 -
Scientific Reports May 2017Anaerobic digesters are man-made habitats for fermentative and methanogenic microbes, and are characterized by extremely high concentrations of organics. However, little...
Anaerobic digesters are man-made habitats for fermentative and methanogenic microbes, and are characterized by extremely high concentrations of organics. However, little is known about how microbes adapt to such habitats. In the present study, we report phylogenetic, metagenomic, and metatranscriptomic analyses of microbiomes in thermophilic packed-bed digesters fed acetate as the major substrate, and we have shown that acetoclastic and hydrogenotrophic methanogens that utilize acetate as a carbon source dominate there. Deep sequencing and precise binning of the metagenomes reconstructed complete genomes for two dominant methanogens affiliated with the genera Methanosarcina and Methanothermobacter, along with 37 draft genomes. The reconstructed Methanosarcina genome was almost identical to that of a thermophilic acetoclastic methanogen Methanosarcina thermophila TM-1, indicating its cosmopolitan distribution in thermophilic digesters. The reconstructed Methanothermobacter (designated as Met2) was closely related to Methanothermobacter tenebrarum, a non-autotrophic hydrogenotrophic methanogen that grows in the presence of acetate. Met2 lacks the Cdh complex required for CO fixation, suggesting that it requires organic molecules, such as acetate, as carbon sources. Although the metagenomic analysis also detected autotrophic methanogens, they were less than 1% in abundance of Met2. These results suggested that non-autotrophic methanogens preferentially grow in anaerobic digesters containing high concentrations of organics.
Topics: Anaerobiosis; Autotrophic Processes; Bioreactors; Contig Mapping; Genes, Archaeal; Methane; Methanobacteriaceae; Methanosarcina; Phylogeny; RNA, Ribosomal, 16S
PubMed: 28473726
DOI: 10.1038/s41598-017-01752-x -
Applied Microbiology and Biotechnology Sep 2018Accumulation of carbon dioxide (CO), associated with global temperature rise, and drastically decreasing fossil fuels necessitate the development of improved renewable...
Accumulation of carbon dioxide (CO), associated with global temperature rise, and drastically decreasing fossil fuels necessitate the development of improved renewable and sustainable energy production processes. A possible route for CO recycling is to employ autotrophic and hydrogenotrophic methanogens for CO-based biological methane (CH) production (CO-BMP). In this study, the physiology and productivity of Methanobacterium thermaggregans was investigated in fed-batch cultivation mode. It is shown that M. thermaggregans can be reproducibly adapted to high agitation speeds for an improved CH productivity. Moreover, inoculum size, sulfide feeding, pH, and temperature were optimized. Optimization of growth and CH productivity revealed that M. thermaggregans is a slightly alkaliphilic and thermophilic methanogen. Hitherto, it was only possible to grow seven autotrophic, hydrogenotrophic methanogenic strains in fed-batch cultivation mode. Here, we show that after a series of optimization and growth improvement attempts another methanogen, M. thermaggregas could be adapted to be grown in fed-batch cultivation mode to cell densities of up to 1.56 g L. Moreover, the CH evolution rate (MER) of M. thermaggregans was compared to Methanothermobacter marburgensis, the CO-BMP model organism. Under optimized cultivation conditions, a maximum MER of 96.1 ± 10.9 mmol L h was obtained with M. thermaggregans-97% of the maximum MER that was obtained utilizing M. marburgensis in a reference experiment. Therefore, M. thermaggregans can be regarded as a CH cell factory highly suited to be applicable for CO-BMP.
Topics: Bioreactors; Carbon Dioxide; Methane; Methanobacterium
PubMed: 29959465
DOI: 10.1007/s00253-018-9183-2 -
The ISME Journal Nov 2023Propionate is a key intermediate in anaerobic digestion processes and often accumulates in association with perturbations, such as elevated levels of ammonia. Under such...
Propionate is a key intermediate in anaerobic digestion processes and often accumulates in association with perturbations, such as elevated levels of ammonia. Under such conditions, syntrophic ammonia-tolerant microorganisms play a key role in propionate degradation. Despite their importance, little is known about these syntrophic microorganisms and their cross-species interactions. Here, we present metagenomes and metatranscriptomic data for novel thermophilic and ammonia-tolerant syntrophic bacteria and the partner methanogens enriched in propionate-fed reactors. A metagenome for a novel bacterium for which we propose the provisional name 'Candidatus Thermosyntrophopropionicum ammoniitolerans' was recovered, together with mapping of its highly expressed methylmalonyl-CoA pathway for syntrophic propionate degradation. Acetate was degraded by a novel thermophilic syntrophic acetate-oxidising candidate bacterium. Electron removal associated with syntrophic propionate and acetate oxidation was mediated by the hydrogen/formate-utilising methanogens Methanoculleus sp. and Methanothermobacter sp., with the latter observed to be critical for efficient propionate degradation. Similar dependence on Methanothermobacter was not seen for acetate degradation. Expression-based analyses indicated use of both H and formate for electron transfer, including cross-species reciprocation with sulphuric compounds and microbial nanotube-mediated interspecies interactions. Batch cultivation demonstrated degradation rates of up to 0.16 g propionate L day at hydrogen partial pressure 4-30 Pa and available energy was around -20 mol propionate. These observations outline the multiple syntrophic interactions required for propionate oxidation and represent a first step in increasing knowledge of acid accumulation in high-ammonia biogas production systems.
Topics: Propionates; Ammonia; Anaerobiosis; Bacteria; Acetates; Methanobacteriaceae; Euryarchaeota; Formates; Hydrogen; Methane
PubMed: 37679429
DOI: 10.1038/s41396-023-01504-y -
Nature Chemical Biology May 2021YcaO enzymes catalyze several post-translational modifications on peptide substrates, including thioamidation, which substitutes an amide oxygen with sulfur. Most...
YcaO enzymes catalyze several post-translational modifications on peptide substrates, including thioamidation, which substitutes an amide oxygen with sulfur. Most predicted thioamide-forming YcaO enzymes are encoded adjacent to TfuA, which when present, is required for thioamidation. While activation of the peptide amide backbone is well established for YcaO enzymes, the function of TfuA has remained enigmatic. Here we characterize the TfuA protein involved in methyl-coenzyme M reductase thioamidation and demonstrate that TfuA catalyzes the hydrolysis of thiocarboxylated ThiS (ThiS-COSH), a proteinaceous sulfur donor, and enhances the affinity of YcaO toward the thioamidation substrate. We also report a crystal structure of a TfuA, which displays a new protein fold. Our structural and mutational analyses of TfuA have uncovered conserved binding interfaces with YcaO and ThiS in addition to revealing a hydrolase-like active site featuring a Ser-Lys catalytic pair.
Topics: Archaeal Proteins; Binding Sites; Cloning, Molecular; Crystallography, X-Ray; Escherichia coli; Euryarchaeota; Gene Expression; Genetic Vectors; Histidine; Kinetics; Mannose-Binding Lectin; Methanobacteriaceae; Methanocaldococcus; Models, Molecular; Mutation; Oligopeptides; Oxidoreductases; Peptides; Protein Binding; Protein Conformation, alpha-Helical; Protein Conformation, beta-Strand; Protein Interaction Domains and Motifs; Protein Processing, Post-Translational; Recombinant Fusion Proteins; Substrate Specificity; Thioamides
PubMed: 33707784
DOI: 10.1038/s41589-021-00771-0