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Applied and Environmental Microbiology Aug 2022The marine bacterium Alcanivorax borkumensis produces a surface-active glycine-glucolipid during growth with long-chain alkanes. A high-performance liquid chromatography...
The marine bacterium Alcanivorax borkumensis produces a surface-active glycine-glucolipid during growth with long-chain alkanes. A high-performance liquid chromatography (HPLC) method was developed for absolute quantification. This method is based on the conversion of the glycine-glucolipid to phenacyl esters with subsequent measurement by HPLC with diode array detection (HPLC-DAD). Different molecular species were separated by HPLC and identified as glucosyl-tetra(3-hydroxy-acyl)-glycine with varying numbers of 3-hydroxy-decanoic acid or 3-hydroxy-octanoic acid groups via mass spectrometry. The growth rate of cells with pyruvate as the sole carbon source was elevated compared to hexadecane as recorded by the increase in cell density as well as oxygen/carbon dioxide transfer rates. The amount of the glycine-glucolipid produced per cell during growth on hexadecane was higher compared with growth on pyruvate. The glycine-glucolipid from pyruvate-grown cells contained considerable amounts of 3-hydroxy-octanoic acid, in contrast to hexadecane-grown cells, which almost exclusively incorporated 3-hydroxy-decanoic acid into the glycine-glucolipid. The predominant proportion of the glycine-glucolipid was found in the cell pellet, while only minute amounts were present in the cell-free supernatant. The glycine-glucolipid isolated from the bacterial cell broth, cell pellet, or cell-free supernatant showed the same structure containing a glycine residue, in contrast to previous reports, which suggested that a glycine-free form of the glucolipid exists which is secreted into the supernatant. In conclusion, the glycine-glucolipid of is resident to the cell wall and enables the bacterium to bind and solubilize alkanes at the lipid-water interface. Alcanivorax borkumensis is one of the most abundant marine bacteria found in areas of oil spills, where it degrades alkanes. The production of a glycine-glucolipid is considered an essential element for alkane degradation. We developed a quantitative method and determined the structure of the glycine-glucolipid in different fractions of the cultures after growth in various media. Our results show that the amount of the glycine-glucolipid in the cells by far exceeds the amount measured in the supernatant, confirming the proposed cell wall localization. These results support the scenario that the surface hydrophobicity of cells increases by producing the glycine-glucolipid, allowing the cells to attach to the alkane-water interface and form a biofilm. We found no evidence for a glycine-free form of the glucolipid.
Topics: Alcanivoraceae; Alkanes; Bacteria; Biodegradation, Environmental; Cell Wall; Glycine; Pyruvic Acid; Water
PubMed: 35938787
DOI: 10.1128/aem.01126-22 -
Sheng Wu Gong Cheng Xue Bao = Chinese... Dec 2021Methanogens are unique microorganisms for methane production and the main contributor of the biogenic methane in atmosphere. Methyl-coenzyme M reductase (Mcr) catalyzes... (Review)
Review
Methanogens are unique microorganisms for methane production and the main contributor of the biogenic methane in atmosphere. Methyl-coenzyme M reductase (Mcr) catalyzes the last step of methane production in methanogenesis and the first step of methane activation in anaerobic oxidation of methane. The genes encoding this enzyme are highly conserved and are widely used as a marker in the identification and phylogenetic study of archaea. There has been a longstanding interest in its unique cofactor F430 and the underpinning mechanisms of enzymatic cleavage of alkane C-H bond. The recent breakthroughs of high-resolution protein and catalytic-transition-state structures further advanced the structure-function study of Mcr. In particular, the recent discovery of methyl-coenzyme M reductase-like (Mcr-like) enzymes that activates the anaerobic degradation of non-methane alkanes has attracted much interest in the molecular mechanisms of C-H activation without oxygen. This review summarized the advances on function-structure-mechanism study of Mcr/Mcr-like enzymes. Additionally, future directions in anaerobic oxidation of alkanes and greenhouse-gas control using Mcr/Mcr-like enzymes were proposed.
Topics: Archaea; Methane; Oxidation-Reduction; Oxidoreductases; Phylogeny
PubMed: 34984864
DOI: 10.13345/j.cjb.200830 -
Metabolic and stress responses of Acinetobacter oleivorans DR1 during long-chain alkane degradation.Microbial Biotechnology Nov 2017Acinetobacter oleivorans DR1 can utilize C -C alkanes as a sole carbon source but not short-chain alkanes (C , C ). Two copies of each alkB-, almA- and ladA-type alkane...
Acinetobacter oleivorans DR1 can utilize C -C alkanes as a sole carbon source but not short-chain alkanes (C , C ). Two copies of each alkB-, almA- and ladA-type alkane hydroxylase (AH) are present in the genome of DR1 cells. Expression and mutational analyses of AHs showed that alkB1 and alkB2 are the major AH-encoding genes under C -C , and the roles of other almA- and ladA genes are negligible. Our data suggested that AlkB1 is responsible for long-chain alkane utilization (C -C ), and AlkB2 is important for medium-chain alkane (C -C ) metabolism. Phylogenetic analyses revealed large incongruities between phylogenies of 16S rRNA and each AH gene, which implies that A. oleivorans DR1 has acquired multiple alkane hydroxylases through horizontal gene transfer. Transcriptomic and qRT-PCR analyses suggested that genes participating in the synthesis of siderophore, trehalose and poly 3-hydroxybutyrate (PHB) were expressed at much higher levels when cells used C than when used succinate as a carbon source. The following biochemical assays supported our gene expression analyses: (i) quantification of siderophore, (ii) measurement of trehalose and (iii) observation of PHB storage. Interestingly, highly induced both ackA gene encoding an acetate kinase A and pta gene encoding a phosphotransacetylase suggested unusual ATP synthesis during C alkane degradation, which was demonstrated by ATP measurement using the ΔackA mutant. Impaired growth of the ΔaceA mutant indicated that the glyoxylate shunt pathway is important when C alkane is utilized. Our data provide insight into long-chain alkane degradation in soil microorganisms.
Topics: Acinetobacter; Alkanes; Bacteria; Bacterial Proteins; Biodegradation, Environmental; Cytochrome P-450 CYP4A; Phylogeny; Siderophores; Trehalose
PubMed: 28857443
DOI: 10.1111/1751-7915.12852 -
Microbiology (Reading, England) Oct 2020Methane-oxidizing bacteria (methanotrophs) play a vital role in reducing atmospheric methane emissions, and hence mitigating their potent global warming effects. A... (Review)
Review
Methane-oxidizing bacteria (methanotrophs) play a vital role in reducing atmospheric methane emissions, and hence mitigating their potent global warming effects. A significant proportion of the methane released is thermogenic natural gas, containing associated short-chain alkanes as well as methane. It was one hundred years following the description of methanotrophs that facultative strains were discovered and validly described. These can use some multi-carbon compounds in addition to methane, often small organic acids, such as acetate, or ethanol, although strains can also use short-chain alkanes, presumably deriving a competitive advantage from this metabolic versatility. Here, we review the diversity and molecular ecology of facultative methanotrophs. We discuss the genetic potential of the known strains and outline the consequent benefits they may obtain. Finally, we review the biotechnological promise of these fascinating microbes.
Topics: Alcohols; Alkanes; Alphaproteobacteria; Bacterial Proteins; Biotechnology; Carbon; Methane; Oxygenases; Phylogeny; Soil Microbiology
PubMed: 33085587
DOI: 10.1099/mic.0.000977 -
Antimicrobial Agents and Chemotherapy Mar 2021Herpesviruses are widespread and can cause serious illness. Many currently available antiviral drugs have limited effects, result in rapid development of resistance, and...
Herpesviruses are widespread and can cause serious illness. Many currently available antiviral drugs have limited effects, result in rapid development of resistance, and often exhibit dose-dependent toxicity. Especially for human cytomegalovirus (HCMV), new well-tolerated compounds with novel mechanisms of action are urgently needed. In this study, we characterized the antiviral activity of two new diazadispiroalkane derivatives, 11826091 and 11826236. These two small molecules exhibited strong activity against low-passage-number HCMV. Pretreatment of cell-free virus with these compounds greatly reduced infection. Time-of-addition assays where 11826091 or 11826236 was added to cells before infection, before and during infection, or during or after infection demonstrated an inhibitory effect on early steps of infection. Interestingly, 11826236 had an effect by addition to cells after infection. Results from entry assays showed the major effect to be on attachment. Only 11826236 had a minimal effect on penetration comparable to heparin. Further, no effect on virus infection was found for cell lines with a defect in heparan sulfate expression or lacking all surface glycosaminoglycans, indicating that these small molecules bind to heparan sulfate on the cell surface. To test this further, we extended our analyses to pseudorabies virus (PrV), a member of the , which is known to use cell surface heparan sulfate for initial attachment via nonessential glycoprotein C (gC). While infection with PrV wild type was strongly impaired by 11826091 or 11826236, as with heparin, a mutant lacking gC was unaffected by either treatment, demonstrating that primary attachment to heparan sulfate via gC is targeted by these small molecules.
Topics: Alkanes; Animals; Antiviral Agents; Glycosaminoglycans; Heparin; Heparitin Sulfate; Herpesvirus 1, Suid; Humans; Spiro Compounds; Viral Envelope Proteins; Virus Internalization
PubMed: 33495228
DOI: 10.1128/AAC.02103-20 -
Journal of Bacteriology Aug 2021Archaeal methanogens, methanotrophs, and alkanotrophs have a high demand for iron (Fe) and sulfur (S); however, little is known of how they acquire, traffic, deploy, and...
Archaeal methanogens, methanotrophs, and alkanotrophs have a high demand for iron (Fe) and sulfur (S); however, little is known of how they acquire, traffic, deploy, and store these elements. Here, we examined the distribution of homologs of proteins mediating key steps in Fe/S metabolism in model microorganisms, including iron(II) sensing/uptake (FeoAB), sulfide extraction from cysteine (SufS), and the biosynthesis of iron-sulfur [Fe-S] clusters (SufBCDE), siroheme (Pch2 dehydrogenase), protoheme (AhbABCD), cytochrome (Cyt ) (CcmCF), and iron storage/detoxification (Bfr, FtrA, and IssA), among 326 publicly available, complete or metagenome-assembled genomes of archaeal methanogens/methanotrophs/alkanotrophs. The results indicate several prevalent but nonuniversal features, including FeoB, SufBC, and the biosynthetic apparatus for the basic tetrapyrrole scaffold, as well as its siroheme (and F) derivatives. However, several early-diverging genomes lacked SufS and pathways to synthesize and deploy heme. Genomes encoding complete versus incomplete heme biosynthetic pathways exhibited equivalent prevalences of [Fe-S] cluster binding proteins, suggesting an expansion of catalytic capabilities rather than substitution of heme for [Fe-S] in the former group. Several strains with heme binding proteins lacked heme biosynthesis capabilities, while other strains with siroheme biosynthesis capability lacked homologs of known siroheme binding proteins, indicating heme auxotrophy and unknown siroheme biochemistry, respectively. While ferritin proteins involved in ferric oxide storage were widespread, those involved in storing Fe as thioferrate were unevenly distributed. Collectively, the results suggest that differences in the mechanisms of Fe and S acquisition, deployment, and storage have accompanied the diversification of methanogens/methanotrophs/alkanotrophs, possibly in response to differential availability of these elements as these organisms evolved. Archaeal methanogens, methanotrophs, and alkanotrophs, argued to be among the most ancient forms of life, have a high demand for iron (Fe) and sulfur (S) for cofactor biosynthesis, among other uses. Here, using comparative bioinformatic approaches applied to 326 genomes, we show that major differences in Fe/S acquisition, trafficking, deployment, and storage exist in this group. Variation in these characters was generally congruent with the phylogenetic placement of these genomes, indicating that variation in Fe/S usage and deployment has contributed to the diversification and ecology of these organisms. However, incongruency was observed among the distribution of cofactor biosynthesis pathways and known protein destinations for those cofactors, suggesting auxotrophy or yet-to-be-discovered pathways for cofactor biosynthesis.
Topics: Alkanes; Archaea; Archaeal Proteins; Autotrophic Processes; Biosynthetic Pathways; Coenzymes; Cysteine; Ferric Compounds; Heme; Iron; Iron-Sulfur Proteins; Methane; Phylogeny; Sulfur
PubMed: 34124941
DOI: 10.1128/JB.00117-21 -
Angewandte Chemie (International Ed. in... May 2022Oxidase-type oxidation is an attractive strategy in organic synthesis due to the use of O as the terminal oxidant. Organic photocatalysis can effect metal-free oxidase...
Oxidase-type oxidation is an attractive strategy in organic synthesis due to the use of O as the terminal oxidant. Organic photocatalysis can effect metal-free oxidase chemistry. Nevertheless, current methods are limited in reaction scope, possibly due to the lack of suitable photocatalysts. Here we report an isoquinoline-derived diaryl ketone-type photocatalyst, which has much enhanced absorption of blue and visible light compared to conventional diaryl ketones. This photocatalyst enables dehydrogenative cross-coupling of heteroarenes with unactivated and activated alkanes as well as aldehydes using air as the oxidant. A wide range of heterocycles with various functional groups are suitable substrates. Transient absorption and excited-state quenching experiments point to an unconventional mechanism that involves an excited state "self-quenching" process to generate the N-radical cation form of the sensitizer, which subsequently abstracts a hydrogen atom from the alkane substrate to yield a reactive alkyl radical.
Topics: Alkanes; Catalysis; Hydrogen; Isoquinolines; Ketones; Oxidants; Oxidoreductases
PubMed: 35253971
DOI: 10.1002/anie.202202649 -
Nature Microbiology Feb 2023'Candidatus Methanophagales' (ANME-1) is an order-level clade of archaea responsible for anaerobic methane oxidation in deep-sea sediments. The diversity, ecology and...
'Candidatus Methanophagales' (ANME-1) is an order-level clade of archaea responsible for anaerobic methane oxidation in deep-sea sediments. The diversity, ecology and evolution of ANME-1 remain poorly understood. In this study, we use metagenomics on deep-sea hydrothermal samples to expand ANME-1 diversity and uncover the effect of virus-host dynamics. Phylogenetic analyses reveal a deep-branching, thermophilic family, 'Candidatus Methanospirareceae', closely related to short-chain alkane oxidizers. Global phylogeny and near-complete genomes show that hydrogen metabolism within ANME-1 is an ancient trait that was vertically inherited but differentially lost during lineage diversification. Metagenomics also uncovered 16 undescribed virus families so far exclusively targeting ANME-1 archaea, showing unique structural and replicative signatures. The expansive ANME-1 virome contains a metabolic gene repertoire that can influence host ecology and evolution through virus-mediated gene displacement. Our results suggest an evolutionary continuum between anaerobic methane and short-chain alkane oxidizers and underscore the effects of viruses on the dynamics and evolution of methane-driven ecosystems.
Topics: Archaea; Ecosystem; Phylogeny; Virome; Geologic Sediments; Anaerobiosis; Methane; Alkanes
PubMed: 36658397
DOI: 10.1038/s41564-022-01297-4 -
Spectrochimica Acta. Part A, Molecular... Nov 2022n-Alkanes and long-chain alcohols (LCOH) have been used as faecal markers to assess the feeding behaviour of both wild and domestic herbivore species. However, their...
Application of Fourier transform infrared spectroscopy (FTIR) techniques in the mid-IR (MIR) and near-IR (NIR) spectroscopy to determine n-alkane and long-chain alcohol contents in plant species and faecal samples.
n-Alkanes and long-chain alcohols (LCOH) have been used as faecal markers to assess the feeding behaviour of both wild and domestic herbivore species. However, their chemical analysis is time-consuming and expensive, making it necessary to develop more expeditious methodologies to evaluate concentrations of these markers. This work aimed to evaluate the use of Fourier Transform Infrared Spectroscopy (FTIR) technology in the near infrared (NIR) and mid infrared (MIR) intervals, for the determination of n-alkane and LCOH concentrations of different plant species and faecal samples of domestic herbivores. Spectra of 33 feed samples, namely L. perenne, T. repens, U. gallii, short heathers (mixture of Erica spp. and Calluna vulgaris), improved pasture grasses (mixture of L. perenne and A. capillaris), heath grasses (mixture of P. longifolium and A. curtissii), improved pasture species (mixture of L. perenne, T. repens and A. capillaris) and herbaceous species (mixture of all herbaceous species found in the plot)) and 181 faecal samples (cattle and horses) were recorded. In order to develop calibrations for the prediction of n-alkanes and LCOH concentrations, partial least squares (PLS) regression was used. Regarding the models developed for plant species, the best results were observed for the calibrations using NIR. The best external validation coefficients of determination (Rv) obtained were 0.90 and 0.79 for LCOH and n-alkanes, respectively. For faecal samples, in the NIR interval, results indicate similar external validation predictions (Rv) for both animal species (0.64). On the contrary, in the MIR interval, differences between cattle (0.70) and horses (0.57) faecal samples in Rv were observed. Regarding the models created for both animal species faeces, LCOH (C-OH and C-OH concentrations ranging from 713.3 to 4451.9 mg/kg DM, respectively; Rv values ranging from 0.72 to 0.95) and n-alkanes (C31 and C33 concentrations ranging from 112.8 to 643.2 mg/kg DM, respectively; Rv values ranging from 0.19 to 0.90) present in higher concentrations tended to be those with better estimates. Results obtained suggest that the selection of the technique to be used may depend on the type of matrix, being the homogeneity of the matrices one of the most important factors for its success. In order to improve the accuracy and robustness of the models created for the estimation of the concentrations of these markers using these methodologies, the database (greater variability) used for the calibrations of these models must be increased.
Topics: Alkanes; Animals; Biomarkers; Cattle; Feces; Feeding Behavior; Horses; Plants; Spectroscopy, Fourier Transform Infrared
PubMed: 35753098
DOI: 10.1016/j.saa.2022.121544 -
Journal of Hazardous Materials Apr 2022Cyclic alkanes (c-alkanes) are toxic compounds that are abundant in subsurface oil reservoirs and spilled condensate; hence, their environmental risk is significant....
Cyclic alkanes (c-alkanes) are toxic compounds that are abundant in subsurface oil reservoirs and spilled condensate; hence, their environmental risk is significant. Although numerous studies have focused on the decomposition of other compound classes, e.g., acyclic alkanes and aromatic hydrocarbons, very little is known about the biodegradation of c-alkanes in the marine environment. Here, we enriched methylcyclohexane (MCH)-degrading bacteria derived from the cold bottom water (10-20 °C) of China's marginal seas in summer and characterized the changes to the bacterial community using high-throughput amplicon sequencing. MCH-consuming bacteria failed to grow from the warmer surface water (25-29 °C) in the same geographic sites and seasons. Notably, MCH-consuming communities derived from the cold bottom water in the Yellow Sea exhibit distinct structures compared to the other treatments. Furthermore, almost all dominant species in this setting appear to be specifically adapted to deeper cold water as indicated by significantly negative correlations to temperature (P < 0.01). From these results, we proposed that the biodegradation of MCH is effectively limited to the colder waters (10-20 °C) of China's marginal seas, with uncultured psychrophiles acting as the key taxa for MCH decomposition. Overall, this study indicates key functions for uncultivated microbes in the marine environment.
Topics: Alkanes; Bacteria; Biodegradation, Environmental; China; Oceans and Seas; Seawater
PubMed: 34991007
DOI: 10.1016/j.jhazmat.2021.128129