The nickel enzyme methyl-coenzyme M reductase from methanogenic archaea: in vitro interconversions among the EPR detectable MCR-red1 and MCR-red2 states

2001 ◽  
Vol 7 (1-2) ◽  
pp. 101-112 ◽  
Author(s):  
Felix Mahlert ◽  
Wolfgang Grabarse ◽  
Jörg Kahnt ◽  
Rudolf K. Thauer ◽  
Evert C. Duin
Keyword(s):  
Methanogenic Archaea ◽  
Coenzyme M ◽  
Nickel Enzyme ◽  
Methyl Coenzyme M Reductase

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.

scholarly journals Mode of action uncovered for the specific reduction of methane emissions from ruminants by the small molecule 3-nitrooxypropanol

2016 ◽  
Vol 113 (22) ◽  
pp. 6172-6177 ◽  
Author(s):  
Evert C. Duin ◽  
Tristan Wagner ◽  
Seigo Shima ◽  
Divya Prakash ◽  
Bryan Cronin ◽  
...  
Keyword(s):  
Active Site ◽  
Energy Content ◽  
Methanogenic Archaea ◽  
Methane Emissions ◽  
In Vivo Experiments ◽  
Nickel Enzyme ◽  
Pure Cultures ◽  
Negative Side Effects

Ruminants, such as cows, sheep, and goats, predominantly ferment in their rumen plant material to acetate, propionate, butyrate, CO2, and methane. Whereas the short fatty acids are absorbed and metabolized by the animals, the greenhouse gas methane escapes via eructation and breathing of the animals into the atmosphere. Along with the methane, up to 12% of the gross energy content of the feedstock is lost. Therefore, our recent report has raised interest in 3-nitrooxypropanol (3-NOP), which when added to the feed of ruminants in milligram amounts persistently reduces enteric methane emissions from livestock without apparent negative side effects [Hristov AN, et al. (2015)Proc Natl Acad Sci USA112(34):10663–10668]. We now show with the aid of in silico, in vitro, and in vivo experiments that 3-NOP specifically targets methyl-coenzyme M reductase (MCR). The nickel enzyme, which is only active when its Ni ion is in the +1 oxidation state, catalyzes the methane-forming step in the rumen fermentation. Molecular docking suggested that 3-NOP preferably binds into the active site of MCR in a pose that places its reducible nitrate group in electron transfer distance to Ni(I). With purified MCR, we found that 3-NOP indeed inactivates MCR at micromolar concentrations by oxidation of its active site Ni(I). Concomitantly, the nitrate ester is reduced to nitrite, which also inactivates MCR at micromolar concentrations by oxidation of Ni(I). Using pure cultures, 3-NOP is demonstrated to inhibit growth of methanogenic archaea at concentrations that do not affect the growth of nonmethanogenic bacteria in the rumen.

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 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.

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