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At least 109 records · Page 6
All sourcesNASA NTRSDOE OSTI
DOE OSTI2023

Diverse electron carriers drive syntrophic interactions in an enriched anaerobic acetate-oxidizing consortium

Abstract In many anoxic environments, syntrophic acetate oxidation (SAO) is a key pathway mediating the conversion of acetate into methane through obligate cross-feeding interactions between SAO bacteria (SAOB) and methanogenic archaea. The SAO pathway is particularly important in engineered environments such as anaerobic digestion (AD) systems operating at thermophilic temperatures and/or with high ammonia. Despite the widespread importance of SAOB to the stability of the AD process, little is known about their in situ physiologies due to typically low biomass yields and resistance to isolation. Here, we performed a long-term (300-day) continuous enrichment of a thermophilic (55 °C) SAO community from a municipal AD system using acetate as the sole carbon source. Over 80% of the enriched bioreactor metagenome belonged to a three-member consortium, including an acetate-oxidizing bacterium affiliated with DTU068 encoding for carbon dioxide, hydrogen, and formate production, along with two methanogenic archaea affiliated with Methanothermobacter_A. Stable isotope probing was coupled with metaproteogenomics to quantify carbon flux into each community member during acetate conversion and inform metabolic reconstruction and genome-scale modeling. This effort revealed that the two Methanothermobacter_A species differed in their preferred electron donors, with one possessing the ability to grow on formate and the other only consuming hydrogen. A thermodynamic analysis suggested that the presence of the formate-consuming methanogen broadened the environmental conditions where ATP production from SAO was favorable. Collectively, these results highlight how flexibility in electron partitioning during SAO likely governs community structure and fitness through thermodynamic-driven mutualism, shedding valuable insights into the metabolic underpinnings of this key functional group within methanogenic ecosystems.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2026

Carbon monoxide oxidation expands the known metabolic capacity in anaerobic methanotrophic consortia

Consortia of anaerobic methane-oxidizing archaea (ANME-2) and sulphate-reducing bacteria (SRB) represent globally relevant syntrophic associations capable of growing with minimal amounts of free energy and can persist when methane becomes limiting. Carbon monoxide (CO) has been reported in seep environments and represents a thermodynamically favourable alternative electron donor due to its low reduction potential. Here, we show that environmental ANME-SRB consortia can oxidize CO in the absence of methane, in anoxic microcosm experiments using a combination of stable isotope geochemical tracers, metatranscriptomics, and single cell activity measurements (FISH–nanoSIMS). The oxidation of CO was coupled with sulphate-reduction by syntrophic consortia, and, in the absence of sulphate, through CO 2 reduction to methane by ANME-2. Under these conditions, the production of methane was one ninth the rate of methanotrophy coupled to sulphate-reduction. Paired single cell FISH-nanoSIMS analysis of anabolic activity indicates that CO respiration appears to support cell maintenance rather than active growth, consistent with the observed down-regulation of energy generating complexes in ANME (e.g., mtr, rnf, etc.). The versatile capability of CO oxidation by anaerobic methanotrophic consortia broadens our understanding of carbon cycling in methane seeps and highlights potential mechanisms of resilience by methanotrophic archaea under changing geochemical regimes.

03 NATURAL GAS↗
DOE OSTI2022

Biogeochemical Characteristics of Earth's Volcanic Permafrost: An Analog of Extraterrestrial Environments

This article describes a study of frozen volcanic deposits collected from volcanoes Tolbachik and Bezymianny on the Kamchatka Peninsula, Russia, and Deception Island volcano, Antarctica. In addition, we studied suprasnow ash layers deposited after the 2007 eruptions of volcanoes Shiveluch and Bezymianny on Kamchatka. The main objectives were to characterize the presence and survivability of thermophilic microorganisms in perennially frozen volcanic deposits. As opposed to permafrost from the polar regions, viable thermophiles were detected in volcanic permafrost by cultivation, microscopy, and sequencing. In the permafrost of Tolbachik volcano, we observed methane formation by both psychrophilic and thermophilic methanogenic archaea, while at 37°C, methane production was noticeably lower. Thermophilic bacteria isolated from volcanic permafrost from the Deception Island were 99.93% related to Geobacillus stearothermophilus. Furthermore, our data showed biological sulfur reduction to sulfide at 85°C and even at 130°C, where hyperthermophilic archaea of the genus Thermoproteus were registered. Sequences of hyperthermophilic bacteria of the genus Caldicellulosiruptor were discovered in clone libraries from fresh volcanic ash deposited on snow. Microorganisms found in volcanic terrestrial permafrost may serve as a model for the alien inhabitants of Mars, a cryogenic planet with numerous volcanoes. Thermophiles and hyperthermophiles and their metabolic processes represent a guideline for the future exploration missions on Mars.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2020

Kinetic and substrate complex characterization of RamA, a corrinoid protein reductive activase from Methanosarcina barkeri

ABSTRACT In microbial corrinoid-dependent methyltransferase systems, adventitious Co(I)-corrinoid oxidation halts catalysis and necessitates repair by ATP-dependent reductive activases. RamA, an activase with a C-terminal ferredoxin domain with two [4Fe-4S] clusters from methanogenic archaea, has been far less studied than the bacterial activases bearing an N-terminal ferredoxin domain with one [2Fe-2S] cluster. These differences suggest RamA might prove to have other distinctive characteristics. Here, we examine RamA kinetics and the stoichiometry of the corrinoid protein:RamA complex. Like bacterial activases, K+ stimulates RamA. Potassium stimulation had been questioned due to differences in the primary structure of bacterial and methanogen activases. Unlike one bacterial activase, ATP is not inhibitory allowing the first determination of apparent kinetic parameters for any corrinoid activase. Unlike bacterial activases, a single RamA monomer complexes a single corrinoid protein monomer. Alanine replacement of a RamA serine residue corresponding to the serine of one bacterial activase which ligates the corrinoid cobalt during complex formation led to only moderate changes in the kinetics of RamA. These results reveal new differences in the two types of corrinoid activases, and provide direct evidence for the proposal that corrinoid activases act as catalytic monomers, unlike other enzymes that couple ATP hydrolysis to difficult reductions.

Microbiology↗
DOE OSTI2021

Beyond the usual suspects: methanogenic communities in eastern North American peatlands are also influenced by nickel and copper concentrations

Peatlands both accumulate carbon and release methane, but their broad range in environmental conditions means that the diversity of microorganisms responsible for carbon cycling is still uncertain. Here, we describe a community analysis of methanogenic archaea responsible for methane production in 17 peatlands from 36 to 53 N latitude across the eastern half of North America, including three metal-contaminated sites. Methanogenic community structure was analysed through Illumina amplicon sequencing of the mcrA gene. Whether metal-contaminated sites were included or not, metal concentrations in peat were a primary driver of methanogenic community composition, particularly nickel, a trace element required in the F 430 cofactor in methyl-coenzyme M reductase that is also toxic at high concentrations. Copper was also a strong predictor, likely due to inhibition at toxic levels and/or to cooccurrence with nickel, since copper enzymes are not known to be present in anaerobic archaea. Here, the methanogenic groups Methanocellales and Methanosarcinales were prevalent in peatlands with low nickel concentrations, while Methanomicrobiales and Methanomassiliicoccales were abundant in peatlands with higher nickel concentrations. Results suggest that peat-associated trace metals are predictors of methanogenic communities in peatlands.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2023

Structure, substrate binding and activity of a unique AAA+ protein: the BrxL phage restriction factor

Abstract Bacteriophage exclusion (‘BREX’) systems are multi-protein complexes encoded by a variety of bacteria and archaea that restrict phage by an unknown mechanism. One BREX factor, termed BrxL, has been noted to display sequence similarity to various AAA+ protein factors including Lon protease. In this study we describe multiple CryoEM structures of BrxL that demonstrate it to be a chambered, ATP-dependent DNA binding protein. The largest BrxL assemblage corresponds to a dimer of heptamers in the absence of bound DNA, versus a dimer of hexamers when DNA is bound in its central pore. The protein displays DNA-dependent ATPase activity, and ATP binding promotes assembly of the complex on DNA. Point mutations within several regions of the protein-DNA complex alter one or more in vitro behaviors and activities, including ATPase activity and ATP-dependent association with DNA. However, only the disruption of the ATPase active site fully eliminates phage restriction, indicating that other mutations can still complement BrxL function within the context of an otherwise intact BREX system. BrxL displays significant structural homology to MCM subunits (the replicative helicase in archaea and eukaryotes), implying that it and other BREX factors may collaborate to disrupt initiation of phage DNA replication.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2023

Structural basis for the tRNA-dependent activation of the terminal complex of selenocysteine synthesis in humans

Abstract O-Phosphoseryl-tRNASec selenium transferase (SepSecS) catalyzes the terminal step of selenocysteine (Sec) synthesis in archaea and eukaryotes. How the Sec synthetic machinery recognizes and discriminates tRNASec from the tRNA pool is essential to the integrity of the selenoproteome. Previously, we suggested that SepSecS adopts a competent conformation that is pre-ordered for catalysis. Herein, using high-resolution X-ray crystallography, we visualized tRNA-dependent conformational changes in human SepSecS that may be a prerequisite for achieving catalytic competency. We show that tRNASec binding organizes the active sites of the catalytic protomer, while stabilizing the N- and C-termini of the non-catalytic protomer. Binding of large anions to the catalytic groove may further optimize the catalytic site for substrate binding and catalysis. Our biochemical and mutational analyses demonstrate that productive SepSecS•tRNASec complex formation is enthalpically driven and primarily governed by electrostatic interactions between the acceptor-, TΨC-, and variable arms of tRNASec and helices α1 and α14 of SepSecS. The detailed visualization of the tRNA-dependent activation of SepSecS provides a structural basis for a revised model of the terminal reaction of Sec formation in archaea and eukaryotes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

Four Families of Folate-independent Methionine Synthases

Abstract Although most organisms synthesize methionine from homocysteine and methyl folates, some have “core” methionine synthases that lack folate-binding domains and use other methyl donors. In vitro , the characterized core synthases use methylcobalamin as a methyl donor, but in vivo , they probably rely on corrinoid (vitamin B12-binding) proteins. We identified four families of core methionine synthases that are distantly related to each other (under 30% pairwise amino acid identity). From the characterized enzymes, we identified the families MesA, which is found in methanogens, and MesB, which is found in anaerobic bacteria and archaea with the Wood-Ljungdahl pathway. A third uncharacterized family, MesC, is found in anaerobic archaea that have the Wood-Ljungdahl pathway and lack known forms of methionine synthase. We predict that most members of the MesB and MesC families accept methyl groups from the iron-sulfur corrinoid protein of that pathway. The fourth family, MesD, is found only in aerobic bacteria. Using transposon mutants and complementation, we show that MesD does not require 5-methyltetrahydrofolate or cobalamin. Instead, MesD requires an uncharacterized protein family (DUF1852) and oxygen for activity.

Price, Morgan↗
DOE OSTI2022

Alterations in soil pH emerge as a key driver of the impact of global change on soil microbial nitrogen cycling: Evidence from a global meta-analysis

Soil nitrogen (N) cycling is critical to the productivity of terrestrial ecosystems. However, the impact of global change factors (GCFs) on the microbial mediators of N cycling pathways has yet to be synthesized, and it also remains unclear whether the response of the abundance of N-cycling genes can predict changes in their corresponding processes. We synthesized 8322 paired observations of soil microorganisms related to N cycling from field experiments in which GCFs (climate change and nutrient addition) were manipulated. We found that the abundance of soil microbes and most N-cycling genes were resistant to elevated CO 2 , experimental warming and water addition/reduction; however, N addition and the combination of N addition with other GCFs significantly increased the abundance of ammonia oxidizer bacteria (amoA-AOB). The results indicated that in steady-state (natural) conditions, the main factors driving the global abundance of soil bacteria, archaea and N-cycling genes varied in terms of the contributions of climatic and edaphic factors. However, upon manipulation of GCFs, the induced change in soil pH was the most essential factor associated with changes in the abundance of soil microbes and N-cycling genes. Notably, the changes in ammonia-oxidizing archaea (amoA-AOA) and amoA-AOB genes, in addition to genes involved in denitrification (nirS and nirK), were significantly correlated with the rates of their corresponding processes, but GCF-induced shifts in the potential nitrification rate (PNR) were explained well by changes in the abundance of the amoA-AOB gene under GCFs. In conclusion, our study highlights how ongoing GCFs impact the abundance of soil microbes and N-cycling genes, which might have a profound impact on terrestrial N cycling. Our field-based results provide new insights into the drivers of the abundance of soil microbes and N-cycling genes.

54 ENVIRONMENTAL SCIENCES↗
DOE OSTI2024

Tetraether archaeal lipids promote long‐term survival in extreme conditions

Abstract The sole unifying feature of the incredibly diverse Archaea is their isoprenoid‐based ether‐linked lipid membranes. Unique lipid membrane composition, including an abundance of membrane‐spanning tetraether lipids, impart resistance to extreme conditions. Many questions remain, however, regarding the synthesis and modification of tetraether lipids and how dynamic changes to archaeal lipid membrane composition support hyperthermophily. Tetraether membranes, termed glycerol dibiphytanyl glycerol tetraethers (GDGTs), are generated by tetraether synthase (Tes) by joining the tails of two bilayer lipids known as archaeol. GDGTs are often further specialized through the addition of cyclopentane rings by GDGT ring synthase (Grs). A positive correlation between relative GDGT abundance and entry into stationary phase growth has been observed, but the physiological impact of inhibiting GDGT synthesis has not previously been reported. Here, we demonstrate that the model hyperthermophile Thermococcus kodakarensis remains viable when Tes (TK2145) or Grs (TK0167) are deleted, permitting phenotypic and lipid analyses at different temperatures. The absence of cyclopentane rings in GDGTs does not impact growth in T. kodakarensis , but an overabundance of rings due to ectopic Grs expression is highly fitness negative at supra‐optimal temperatures. In contrast, deletion of Tes resulted in the loss of all GDGTs, cyclization of archaeol, and loss of viability upon transition to the stationary phase in this model archaea. These results demonstrate the critical roles of highly specialized, dynamic, isoprenoid‐based lipid membranes for archaeal survival at high temperatures.

Biochemistry & Molecular Biology↗
DOE OSTI2024

Arbuscular mycorrhiza convey significant plant carbon to a diverse hyphosphere microbial food web and mineral‐associated organic matter

Summary Arbuscular mycorrhizal fungi (AMF) transport substantial plant carbon (C) that serves as a substrate for soil organisms, a precursor of soil organic matter (SOM), and a driver of soil microbial dynamics. Using two‐chamber microcosms where an air gap isolated AMF from roots, we 13 CO 2 ‐labeled Avena barbata for 6 wk and measured the C Rhizophagus intraradices transferred to SOM and hyphosphere microorganisms. NanoSIMS imaging revealed hyphae and roots had similar 13 C enrichment. SOM density fractionation, 13 C NMR, and IRMS showed AMF transferred 0.77 mg C g −1 of soil (increasing total C by 2% relative to non‐mycorrhizal controls); 33% was found in occluded or mineral‐associated pools. In the AMF hyphosphere, there was no overall change in community diversity but 36 bacterial ASVs significantly changed in relative abundance. With stable isotope probing (SIP)‐enabled shotgun sequencing, we found taxa from the Solibacterales, Sphingobacteriales, Myxococcales, and Nitrososphaerales (ammonium oxidizing archaea) were highly enriched in AMF‐imported 13 C (> 20 atom%). Mapping sequences from 13 C‐SIP metagenomes to total ASVs showed at least 92 bacteria and archaea were significantly 13 C‐enriched. Our results illustrate the quantitative and ecological impact of hyphal C transport on the formation of potentially protective SOM pools and microbial roles in the AMF hyphosphere soil food web.

13C SIP↗
DOE OSTI2025

A dynamic protein interactome drives energy conservation and electron flux in Thermococcus kodakarensis

ABSTRACT Life is supported by energy gains fueled by catabolism of a wide range of substrates, each reliant on the selective partitioning of electrons through redox ( red uction and ox idation) reactions. Electron flux through tunable and regulated protein interactions provides dynamic routes for energy conservation, but how electron flux is regulated in vivo , particularly for archaeal metabolisms that support rapid growth at the thermodynamic limits of life, is poorly understood. Identification of bona fide in vivo protein assemblies and how such assemblies dictate the totality of electron flux is critical to our understanding of the regulation imposed on metabolism, energy production, and energy conservation. Here, 25 key proteins in central metabolic redox pathways in the model, genetically accessible, hyperthermophilic archaeon Thermococcus kodakarensis , were purified to reveal an extensive, dynamic, and tightly interconnected network of protein interactions that responds to environmental cues (such as the availability of various reductive sinks) to direct electron flux to maximize energetic gains. Interactions connecting disparate functions suggest many catabolic and anabolic activities occur in spatial proximity in vivo , and while protein complexes have been historically defined under optimal conditions, many of these complexes appear to maintain alternative partnerships in changing conditions. The totality of the results obtained redefines our understanding of in vivo assemblies driving ancient metabolic strategies supporting the growth of modern Archaea. IMPORTANCE Given the potential for rational genetic manipulations of biofuel- and biotech-promising archaea to yield transformative results for major markets, it is a priority to define how the metabolisms of such species are controlled, at least in part, by in vivo protein assemblies, and from such, define routes of energy flux that can be most efficiently altered toward biofuel or biotechnological gains. Proteinaceous electron carriers (PECs, such as ferredoxins) offer the potential for specific protein–protein interactions to coordinate selective reductive flow. Employing the model, genetically accessible, hyperthermophilic archaeon, Thermococcus kodakarensis , we establish the metabolic protein interactome of 25 key redox proteins, revealing that each redox active protein has a dynamic partnership profile, suggesting catabolic and anabolic activities may occur in concert and in temporal and spatial proximity in vivo . These results reveal critical importance in evaluating the newly identified partnerships and their role and utility in providing regulated redox flux in T. kodakarensis .

Williams, Sere A. (ORCID:0000000235509590)↗
DOE OSTI2023

Expression of V-nitrogenase and Fe-nitrogenase in Methanosarcina acetivorans is controlled by molybdenum, fixed nitrogen, and the expression of Mo-nitrogenase

ABSTRACT All nitrogen-fixing bacteria and archaea (diazotrophs) use molybdenum (Mo) nitrogenase to reduce dinitrogen (N 2 ) to ammonia, with some also containing vanadium (V) and iron-only (Fe) nitrogenases that lack Mo. Among diazotrophs, the regulation and usage of the alternative V-nitrogenase and Fe-nitrogenase in methanogens are largely unknown. Methanosarcina acetivorans contains nif , vnf , and anf gene clusters encoding putative Mo-nitrogenase, V-nitrogenase, and Fe-nitrogenase, respectively. This study investigated nitrogenase expression and growth by M. acetivorans in response to fixed nitrogen, Mo/V availability, and CRISPRi repression of the nif , vnf , and/or anf gene clusters. The availability of Mo and V significantly affected growth of M. acetivorans with N 2 but not with NH 4 Cl. M. acetivorans exhibited the fastest growth rate and highest cell yield during growth with N 2 in medium containing Mo, and the slowest growth in medium lacking Mo and V. qPCR analysis revealed the transcription of the nif operon is only moderately affected by depletion of fixed nitrogen and Mo, whereas vnf and anf transcription increased significantly when fixed nitrogen and Mo were depleted, with removal of Mo being key. Immunoblot analysis revealed Mo-nitrogenase is detected when fixed nitrogen is depleted regardless of Mo availability, while V-nitrogenase and Fe-nitrogenase are detected only in the absence of fixed nitrogen and Mo. CRISPRi repression studies revealed that V-nitrogenase and/or Fe-nitrogenase are required for Mo-independent diazotrophy, and unexpectedly that the expression of Mo-nitrogenase is also required. These results reveal that alternative nitrogenase production in M. acetivorans is tightly controlled and dependent on Mo-nitrogenase expression. IMPORTANCE Methanogens and closely related methanotrophs are the only archaea known or predicted to possess nitrogenase. Methanogens play critical roles in both the global biological nitrogen and carbon cycles. Moreover, methanogens are an ancient microbial lineage and nitrogenase likely originated in methanogens. An understanding of the usage and properties of nitrogenases in methanogens can provide new insight into the evolution of nitrogen fixation and aid in the development nitrogenase-based biotechnology. This study provides the first evidence that a methanogen can produce all three forms of nitrogenases, including simultaneously. The results reveal components of Mo-nitrogenase regulate or are needed to produce V-nitrogenase and Fe-nitrogenase in methanogens, a result not seen in bacteria. Overall, this study provides a foundation to understand the assembly, regulation, and activity of the alternative nitrogenases in methanogens.

Biotechnology & Applied Microbiology↗
DOE OSTI2025

Microbial ecology of acidic, biogenic gypsum: community structure and distribution of extremophiles on freshly formed and relict sulfate deposits in a hydrogen sulfide-rich cave

Sulfate minerals are abundant on the Martian surface, and many of these evaporite deposits are thought to have precipitated from acidic fluids. On Earth, gypsum (CaSO 4 •2H 2 O) and other sulfates sometimes form under acidic conditions, so exploring the extremophilic life that occurs in these mineral environments can help evaluate the astrobiological potential of acid sulfate depositional settings. Here, we characterized the microbial communities associated with acidic gypsum deposits in a sulfuric acid cave, where sulfate precipitation is driven by sulfide-oxidizing bacteria and archaea. We used 16S rRNA gene sequencing and cell counts to characterize gypsum-associated microorganisms in freshly formed and relict deposits throughout the cave, to test how microbial community composition and abundance would vary with distance from the sulfidic water table and with the concentration of H 2 S(g) and other gases in the cave atmosphere. We found that actively forming gypsum in the lower cave levels was colonized by low-diversity communities that have few cells compared to other environments in the cave. The most abundant taxa were Acidithiobacillus, Metallibacterium, Mycobacteria, and three different Thermoplasmatales-group archaea, which occupied distinct niches based on proximity to sulfidic streams and the concentration of gases in the cave air. By contrast, deposits in older cave levels had more diverse communities that were distinct from those associated with freshly formed gypsum and likely represent a community reliant on different energy resources. These findings show that acidic sulfate deposits serve as habitats for extremophilic microorganisms and broaden our knowledge of the life associated with terrestrial sulfates.

58 GEOSCIENCES↗
DOE OSTI2020

Lateral Gene Transfer Drives Metabolic Flexibility in the Anaerobic Methane-Oxidizing Archaeal Family Methanoperedenaceae

AOM by microorganisms limits the atmospheric release of the potent greenhouse gas methane and has consequent importance for the global carbon cycle and climate change modeling. While the oxidation of methane coupled to sulfate by consortia of anaerobic methanotrophic (ANME) archaea and bacteria is well documented, several other potential electron acceptors have also been reported to support AOM. In this study, we identify a number of novel respiratory strategies that appear to have been laterally acquired by members of the Methanoperedenaceae , as they are absent from related archaea and other ANME lineages. Expanding the known metabolic potential for members of the Methanoperedenaceae provides important insight into their ecology and suggests their role in linking methane oxidation to several global biogeochemical cycles.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2024

Diverse and unconventional methanogens, methanotrophs, and methylotrophs in metagenome-assembled genomes from subsurface sediments of the Slate River floodplain, Crested Butte, CO, USA

We use metagenome-assembled genomes (MAGs) to understand single-carbon (C1) compound-cycling—particularly methane-cycling—microorganisms in montane riparian floodplain sediments. We generated 1,233 MAGs (>50% completeness and <10% contamination) from 50- to 150-cm depth below the sediment surface capturing the transition between oxic, unsaturated sediments and anoxic, saturated sediments in the Slate River (SR) floodplain (Crested Butte, CO, USA). We recovered genomes of putative methanogens, methanotrophs, and methylotrophs (n = 57). Methanogens, found only in deep, anoxic depths at SR, originate from three different clades (Methanoregulaceae, Methanotrichaceae, and Methanomassiliicoccales), each with a different methanogenesis pathway; putative methanotrophic MAGs originate from within the Archaea (Candidatus Methanoperedens) in anoxic depths and uncultured bacteria (Ca. Binatia) in oxic depths. Genomes for canonical aerobic methanotrophs were not recovered. Ca. Methanoperedens were exceptionally abundant (~1,400× coverage, >50% abundance in the MAG library) in one sample that also contained aceticlastic methanogens, indicating a potential C1/methane-cycling hotspot. Ca. Methylomirabilis MAGs from SR encode pathways for methylotrophy but do not harbor methane monooxygenase or nitrogen reduction genes. Comparative genomic analysis supports that one clade within the Ca. Methylomirabilis genus is not methanotrophic. The genetic potential for methylotrophy was widespread, with over 10% and 19% of SR MAGs encoding a methanol dehydrogenase or substrate-specific methyltransferase, respectively. MAGs from uncultured Thermoplasmata archaea in the Ca. Gimiplasmatales (UBA10834) contain pathways that may allow for anaerobic methylotrophic acetogenesis. Overall, MAGs from SR floodplain sediments reveal a potential for methane production and consumption in the system and a robust potential for methylotrophy.

58 GEOSCIENCES↗
DOE OSTI2024

Transcriptional response of Methanosarcina acetivorans to repression of the energy-conserving methanophenazine: CoM-CoB heterodisulfide reductase enzyme HdrED

ABSTRACT Methane-producing archaea are key organisms in the anaerobic carbon cycle. These organisms, also called methanogens, grow by converting substrate to methane gas in a process called methanogenesis. Previous research showed that the reduction of the terminal electron acceptor is the rate-limiting step in methanogenesis by Methanosarcina acetivorans . In order to gain insight into how the cells sense and respond to the availability of the terminal electron acceptor, we designed an experiment to deplete cells of the essential terminal oxidase enzyme, HdrED. We found that the depletion of HdrED in vivo results in a higher abundance of transcripts for methyltransferases ( mtaC2, mtaB3, mtaC3 ), coenzyme B biosynthesis, C1 metabolism, and pyrimidine compounds. In most cases, these changes were distinct from transcript abundance changes observed during the transition from exponential growth to stationary phase cultures. These data implicate the methylotrophic methanogenesis regulator MsrC (MA4383) in CoM-S-S-CoB heterodisulfide sensing and indicate cells have a specific mechanism to sense intracellular ratio of CoM-S-S-CoB, coenzyme M, and coenzyme B thiols and further suggest transcripts encoding translation and methanogenesis functions are controlled by feed-forward regulation depending on substrate availability. IMPORTANCE Methanosarcina is an emerging model archaeon and synthetic biology platform for the production of renewable energy and sustainable chemicals to reduce dependence on petroleum. Research into metabolic networks and gene regulation in this organism and other methanogens will inform genome-scale metabolic modeling and microbial function prediction in uncultured or non-model anaerobes and archaea. This study suggests methanogens use unknown mechanisms to efficiently couple methanogenesis to gene regulation via CoM-S-S-CoB and ATP availability.

Buan, Nicole R. (ORCID:000000027560973X)↗
DOE OSTI2020

Iron-Only and Vanadium Nitrogenases: Fail-Safe Enzymes or Something More?

The enzyme molybdenum nitrogenase converts atmospheric nitrogen gas to ammonia and is of critical importance for the cycling of nitrogen in the biosphere and for the sustainability of life. Alternative vanadium and iron-only nitrogenases that are homologous to molybdenum nitrogenases are also found in archaea and bacteria, but they have a different transition metal, either vanadium or iron, at their active sites. So far alternative nitrogenases have only been found in microbes that also have molybdenum nitrogenase. They are less widespread than molybdenum nitrogenase in bacteria and archaea, and they are less efficient. The presumption has been that alternative nitrogenases are fail-safe enzymes that are used in situations where molybdenum is limiting. Recent work indicates that vanadium nitrogenase may play a role in the global biological nitrogen cycle and iron-only nitrogenase may contribute products that shape microbial community interactions in nature.

Microbiology↗