scholarly journals Identification of Methyl Coenzyme M Reductase A (mcrA) Genes Associated with Methane-Oxidizing Archaea

2003 ◽  
Vol 69 (9) ◽  
pp. 5483-5491 ◽  
Author(s):  
Steven J. Hallam ◽  
Peter R. Girguis ◽  
Christina M. Preston ◽  
Paul M. Richardson ◽  
Edward F. DeLong
Keyword(s):  
Methane Oxidation ◽  
Evolutionary Relationship ◽  
Methanogenic Archaea ◽  
Small Subunit ◽  
Anaerobic Methane Oxidation ◽  
Small Subunit Rrna ◽  
Genomic Context ◽  
Coenzyme M ◽  
Phylogenetic Groups ◽  
Methyl Coenzyme M Reductase

ABSTRACT Phylogenetic and stable-isotope analyses implicated two methanogen-like archaeal groups, ANME-1 and ANME-2, as key participants in the process of anaerobic methane oxidation. Although nothing is known about anaerobic methane oxidation at the molecular level, the evolutionary relationship between methane-oxidizing archaea (MOA) and methanogenic archaea raises the possibility that MOA have co-opted key elements of the methanogenic pathway, reversing many of its steps to oxidize methane anaerobically. In order to explore this hypothesis, the existence and genomic conservation of methyl coenzyme M reductase (MCR), the enzyme catalyzing the terminal step in methanogenesis, was studied in ANME-1 and ANME-2 archaea isolated from various marine environments. Clone libraries targeting a conserved region of the alpha subunit of MCR (mcrA) were generated and compared from environmental samples, laboratory-incubated microcosms, and fosmid libraries. Four out of five novel mcrA types identified from these sources were associated with ANME-1 or ANME-2 group members. Assignment of mcrA types to specific phylogenetic groups was based on environmental clone recoveries, selective enrichment of specific MOA and mcrA types in a microcosm, phylogenetic congruence between mcrA and small-subunit rRNA tree topologies, and genomic context derived from fosmid sequences. Analysis of the ANME-1 and ANME-2 mcrA sequences suggested the potential for catalytic activity based on conservation of active-site amino acids. These results provide a basis for identifying methanotrophic archaea with mcrA sequences and define a functional genomic link between methanogenic and methanotrophic archaea.

scholarly journals Phylogenetic and Structural Comparisons of the Three Types of Methyl Coenzyme M Reductase from Methanococcales and Methanobacteriales

2017 ◽  
Vol 199 (16) ◽  
Author(s):  
Tristan Wagner ◽  
Carl-Eric Wegner ◽  
Jörg Kahnt ◽  
Ulrich Ermler ◽  
Seigo Shima
Keyword(s):  
Active Site ◽  
Phylogenetic Analyses ◽  
Methanogenic Archaea ◽  
Methane Formation ◽  
Type I ◽  
Type Ii ◽  
Coenzyme M ◽  
Content Type ◽  
Type Iii ◽  
Methyl Coenzyme M Reductase

ABSTRACT The phylogenetically diverse family of methanogenic archaea universally use methyl coenzyme M reductase (MCR) for catalyzing the final methane-forming reaction step of the methanogenic energy metabolism. Some methanogens of the orders Methanobacteriales and Methanococcales contain two isoenzymes. Comprehensive phylogenetic analyses on the basis of all three subunits grouped MCRs from Methanobacteriales and Methanococcales into three distinct types: (i) MCRs from Methanobacteriales, (ii) MCRs from Methanobacteriales and Methanococcales, and (iii) MCRs from Methanococcales. The first and second types contain MCR isoenzymes I and II from Methanothermobacter marburgensis, respectively; therefore, they were designated MCR type I and type II and accordingly; the third one was designated MCR type III. For comparison with the known MCR type I and type II structures, we determined the structure of MCR type III from Methanotorris formicicus and Methanothermococcus thermolithotrophicus. As predicted, the three MCR types revealed highly similar overall structures and virtually identical active site architectures reflecting the chemically challenging mechanism of methane formation. Pronounced differences were found at the protein surface with respect to loop geometries and electrostatic properties, which also involve the entrance of the active-site funnel. In addition, the C-terminal end of the γ-subunit is prolonged by an extra helix after helix γ8 in MCR type II and type III, which is, however, differently arranged in the two MCR types. MCR types I, II, and III share most of the posttranslational modifications which appear to fine-tune the enzymatic catalysis. Interestingly, MCR type III lacks the methyl-cysteine but possesses in subunit α of M. formicicus a 6-hydroxy-tryptophan, which thus far has been found only in the α-amanitin toxin peptide but not in proteins. IMPORTANCE Methyl coenzyme M reductase (MCR) represents a prime target for the mitigation of methane releases. Phylogenetic analyses of MCRs suggested several distinct sequence clusters; those from Methanobacteriales and Methanococcales were subdivided into three types: MCR type I from Methanobacteriales, MCR type II from Methanobacteriales and Methanococcales, and the newly designated MCR type III exclusively from Methanococcales. We determined the first X-ray structures for an MCR type III. Detailed analyses revealed substantial differences between the three types only in the peripheral region. The subtle modifications identified and electrostatic profiles suggested enhanced substrate binding for MCR type III. In addition, MCR type III from Methanotorris formicicus contains 6-hydroxy-tryptophan, a new posttranslational modification that thus far has been found only in the α-amanitin toxin.

A new intra-aerobic metabolism in the nitrite-dependent anaerobic methane-oxidizing bacterium Candidatus ‘Methylomirabilis oxyfera’

2011 ◽  
Vol 39 (1) ◽  
pp. 243-248 ◽  
Author(s):  
Ming L. Wu ◽  
Katharina F. Ettwig ◽  
Mike S.M. Jetten ◽  
Marc Strous ◽  
Jan T. Keltjens ◽  
...  
Keyword(s):  
Nitric Oxide ◽  
Methane Oxidation ◽  
Current Knowledge ◽  
Methanogenic Archaea ◽  
Anaerobic Methane Oxidation ◽  
Oxidation Pathway ◽  
Sulfate Reducing ◽  
Oxidation Of Methane ◽  
Aerobic Methanotrophs ◽  
Reverse Methanogenesis

Biological methane oxidation proceeds either through aerobic or anaerobic pathways. The newly discovered bacterium Candidatus ‘Methylomirabilis oxyfera’ challenges this dichotomy. This bacterium performs anaerobic methane oxidation coupled to denitrification, but does so in a peculiar way. Instead of scavenging oxygen from the environment, like the aerobic methanotrophs, or driving methane oxidation by reverse methanogenesis, like the methanogenic archaea in sulfate-reducing systems, it produces its own supply of oxygen by metabolizing nitrite via nitric oxide into oxygen and dinitrogen gas. The intracellularly produced oxygen is then used for the oxidation of methane by the classical aerobic methane oxidation pathway involving methane mono-oxygenase. The present mini-review summarizes the current knowledge about this process and the micro-organism responsible for it.

scholarly journals Study of Genetic Diversity of Eukaryotic Picoplankton in Different Oceanic Regions by Small-Subunit rRNA Gene Cloning and Sequencing

2001 ◽  
Vol 67 (7) ◽  
pp. 2932-2941 ◽  
Author(s):  
Beatriz Dı́ez ◽  
Carlos Pedr�s-Ali� ◽  
Ramon Massana
Keyword(s):  
Operational Taxonomic Unit ◽  
Small Subunit ◽  
Small Cells ◽  
Rrna Genes ◽  
Rrna Gene ◽  
Polymorphism Analysis ◽  
Small Subunit Rrna ◽  
Phylogenetic Groups ◽  
18S Rrna Genes ◽  
A Minor

ABSTRACT Very small eukaryotic organisms (picoeukaryotes) are fundamental components of marine planktonic systems, often accounting for a significant fraction of the biomass and activity in a system. Their identity, however, has remained elusive, since the small cells lack morphological features for identification. We determined the diversity of marine picoeukaryotes by sequencing cloned 18S rRNA genes in five genetic libraries from North Atlantic, Southern Ocean, and Mediterranean Sea surface waters. Picoplankton were obtained by filter size fractionation, a step that excluded most large eukaryotes and recovered most picoeukaryotes. Genetic libraries of eukaryotic ribosomal DNA were screened by restriction fragment length polymorphism analysis, and at least one clone of each operational taxonomic unit (OTU) was partially sequenced. In general, the phylogenetic diversity in each library was rather great, and each library included many different OTUs and members of very distantly related phylogenetic groups. Of 225 eukaryotic clones, 126 were affiliated with algal classes, especially the Prasinophyceae, the Prymnesiophyceae, the Bacillariophyceae, and the Dinophyceae. A minor fraction (27 clones) was affiliated with clearly heterotrophic organisms, such as ciliates, the chrysomonad Paraphysomonas, cercomonads, and fungi. There were two relatively abundant novel lineages, novel stramenopiles (53 clones) and novel alveolates (19 clones). These lineages are very different from any organism that has been isolated, suggesting that there are previously unknown picoeukaryotes. Prasinophytes and novel stramenopile clones were very abundant in all of the libraries analyzed. These findings underscore the importance of attempts to grow the small eukaryotic plankton in pure culture.

scholarly journals Functional interactions between post-translationally modified amino acids of methyl-coenzyme M reductase in Methanosarcina acetivorans

2019 ◽  
Author(s):  
Dipti D Nayak ◽  
Andi Liu ◽  
Neha Agrawal ◽  
Roy Rodriguez-Carerro ◽  
Shi-Hui Dong ◽  
...  
Keyword(s):  
Amino Acids ◽  
Methanogenic Archaea ◽  
Physical Structure ◽  
Major Influence ◽  
Methanosarcina Acetivorans ◽  
Coenzyme M ◽  
Phenotypic Analysis ◽  
And Function ◽  
Methyl Coenzyme M Reductase

AbstractMethyl-coenzyme M reductase (MCR) plays an important role in mediating global levels of methane by catalyzing a reversible reaction that leads to the production or consumption of this potent greenhouse gas in methanogenic and methanotrophic archaea. In methanogenic archaea, the alpha subunit of MCR (McrA) typically contains four to six post-translationally modified amino acids near the active site. Recent studies have identified genes that install two of these modifications (thioglycine and 5-(S)-methylarginine), yet little is known about the installation and function of the remaining post-translationally modified residues. Here, we provide in vivo evidence that a dedicated SAM-dependent methyltransferase encoded by a gene we designated mcmA is responsible for formation of S-methylcysteine in Methanosarcina acetivorans McrA. Phenotypic analysis of mutants incapable of cysteine methylation suggests that the S-methylcysteine residue plays an important role in adaptation to a mesophilic lifestyle. To examine the interactions between the S-methylcysteine residue and the previously characterized thioglycine, 5-(S)-methylarginine modifications, we generated M. acetivorans mutants lacking the three known modification genes in all possible combinations. Phenotypic analyses revealed complex, physiologically relevant interactions between the modified residues, which alter the thermal stability of MCR in a combinatorial fashion that is not readily predictable from the phenotypes of single mutants. Surprisingly, high-resolution crystal structures of the various unmodified MCRs were indistinguishable from the fully modified enzyme, suggesting that interactions between the post-translationally modified residues do not exert a major influence on the physical structure of the enzyme, but rather serve to fine-tune the activity and efficiency of MCR.

scholarly journals Localization of Methyl-Coenzyme M Reductase as Metabolic Marker for Diverse Methanogenic Archaea

Archaea ◽  
2013 ◽  
Vol 2013 ◽  
pp. 1-7 ◽  
Author(s):  
Christoph Wrede ◽  
Ulrike Walbaum ◽  
Andrea Ducki ◽  
Iris Heieren ◽  
Michael Hoppert
Keyword(s):  
Polyclonal Antibodies ◽  
Methanogenic Archaea ◽  
Growth Conditions ◽  
Laboratory Culture ◽  
Anaerobic Oxidation Of Methane ◽  
Coenzyme M ◽  
Microbial Biofilms ◽  
Oxidation Of Methane ◽  
Metabolic Marker ◽  
Methyl Coenzyme M Reductase

Methyl-Coenzyme M reductase (MCR) as key enzyme for methanogenesis as well as for anaerobic oxidation of methane represents an important metabolic marker for both processes in microbial biofilms. Here, the potential of MCR-specific polyclonal antibodies as metabolic marker in various methanogenic Archaea is shown. For standard growth conditions in laboratory culture, the cytoplasmic localization of the enzyme inMethanothermobacter marburgensis,Methanothermobacter wolfei,Methanococcus maripaludis,Methanosarcina mazei, and in anaerobically methane-oxidizing biofilms is demonstrated. Under growth limiting conditions on nickel-depleted media, at low linear growth of cultures, a fraction of 50–70% of the enzyme was localized close to the cytoplasmic membrane, which implies “facultative” membrane association of the enzyme. This feature may be also useful for assessment of growth-limiting conditions in microbial biofilms.

scholarly journals High-Density Microarray of Small-Subunit Ribosomal DNA Probes

2002 ◽  
Vol 68 (5) ◽  
pp. 2535-2541 ◽  
Author(s):  
Kenneth H. Wilson ◽  
Wendy J. Wilson ◽  
Jennifer L. Radosevich ◽  
Todd Z. DeSantis ◽  
Vijay S. Viswanathan ◽  
...  
Keyword(s):  
Ribosomal Dna ◽  
Bacterial Species ◽  
Small Subunit ◽  
Rrna Genes ◽  
Biological Research ◽  
Ribosomal Database Project ◽  
Small Subunit Rrna ◽  
Phylogenetic Groups ◽  
Chip Technology ◽  
Small Subunit Ribosomal Dna

ABSTRACT Ribosomal DNA sequence analysis, originally conceived as a way to provide a universal phylogeny for life forms, has proven useful in many areas of biological research. Some of the most promising applications of this approach are presently limited by the rate at which sequences can be analyzed. As a step toward overcoming this limitation, we have investigated the use of photolithography chip technology to perform sequence analyses on amplified small-subunit rRNA genes. The GeneChip (Affymetrix Corporation) contained 31,179 20-mer oligonucleotides that were complementary to a subalignment of sequences in the Ribosomal Database Project (RDP) (B. L. Maidak et al., Nucleic Acids Res. 29:173-174, 2001). The chip and standard Affymetrix software were able to correctly match small-subunit ribosomal DNA amplicons with the corresponding sequences in the RDP database for 15 of 17 bacterial species grown in pure culture. When bacteria collected from an air sample were tested, the method compared favorably with cloning and sequencing amplicons in determining the presence of phylogenetic groups. However, the method could not resolve the individual sequences comprising a complex mixed sample. Given these results and the potential for future enhancement of this technology, it may become widely useful.

scholarly journals Biomarker Evidence for Widespread Anaerobic Methane Oxidation in Mediterranean Sediments by a Consortium of Methanogenic Archaea and Bacteria

2000 ◽  
Vol 66 (3) ◽  
pp. 1126-1132 ◽  
Author(s):  
Richard D. Pancost ◽  
Jaap S. Sinninghe Damsté ◽  
Saskia de Lint ◽  
Marc J. E. C. van der Maarel ◽  
Jan C. Gottschal
Keyword(s):  
Methane Oxidation ◽  
Heterotrophic Bacteria ◽  
Methanogenic Archaea ◽  
Anaerobic Methane Oxidation ◽  
Geochemical Data ◽  
Mud Volcanoes ◽  
Terminal Electron Acceptor ◽  
Sulfate Reducing ◽  
Microbiological Process ◽  
Reducing Bacteria

ABSTRACT Although abundant geochemical data indicate that anaerobic methane oxidation occurs in marine sediments, the linkage to specific microorganisms remains unclear. In order to examine processes of methane consumption and oxidation, sediment samples from mud volcanoes at two distinct sites on the Mediterranean Ridge were collected via the submersible Nautile. Geochemical data strongly indicate that methane is oxidized under anaerobic conditions, and compound-specific carbon isotope analyses indicate that this reaction is facilitated by a consortium of archaea and bacteria. Specifically, these methane-rich sediments contain high abundances of methanogen-specific biomarkers that are significantly depleted in13C (δ13C values are as low as −95‰). Biomarkers inferred to derive from sulfate-reducing bacteria and other heterotrophic bacteria are similarly depleted. Consistent with previous work, such depletion can be explained by consumption of13C-depleted methane by methanogens operating in reverse and as part a consortium of organisms in which sulfate serves as the terminal electron acceptor. Moreover, our results indicate that this process is widespread in Mediterranean mud volcanoes and in some localized settings is the predominant microbiological process.

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