Methanogens and the diversity of archaebacteria.

Microbial cycling of volatile organic sulfur compounds (VOSCs), especially dimethyl sulfide (DMS) and methanethiol (MT), is intensively studied because these compounds play an important role in the processes of global warming, acid precipitation, and the global sulfur cycle. VOSC concentrations in freshwater sediments are low due to the balance between the formation and degradation of these compounds. These reactions occur for the greater part at the oxic/anoxic interphase of sediment and water column. In contrast to marine ecosystems, where dimethylsulfoniopropionate is the main precursor of MT and DMS, in freshwater ecosystems, VOSCs are formed mainly by methylation of sulfide and to a lesser extent from the degradation of S-containing amino acids. One of the major routes for DMS and MT formation through sulfide methylation is anaerobic O-demethylation of methoxylated aromatic compounds. Inhibition studies have revealed that the major part of the endogenously produced MT and DMS is degraded anaerobically by methanogens. The major bacterial groups involved in formation and consumption of VOSCs are described.

Microbial cycling of volatile organic sulfur compounds.

The enzyme system catalyzing the formation of methyl-coenzyme M from methanol and coenzyme M in Methanosarcina barkeri is composed of the three different polypeptides MtaA, MtaB and MtaC of which MtaC harbors a corrinoid prosthetic group. The heterologous expression of mtaA and mtaB in Escherichia coli has been described previously. We report here on the overproduction of the apoprotein of MtaC in E. coli, on its reconstitution to the active holoprotein with either cob(II)alamin or methylcob(III)alamin, and on the properties of the reconstituted corrinoid protein. Reconstituted MtaC was found to contain 1 mol bound cobamide/mol. EPR spectroscopic evidence is presented for a His residue as an axial ligand to Co2+of the bound corrinoid. This active-site His was identified by site-directed mutagenesis as His136 in the MtaC sequence that contains four His residues. The reconstituted MtaC, in the cob(I)amide oxidation state, was methylated with methanol in the presence of MtaB and demethylated with coenzyme M in the presence of MtaA. In the presence of both MtaB and MtaA, methyl-coenzyme M was formed from methanol and coenzyme M at specific rates comparable to those determined for the enzyme system purified from M. barkeri. M. barkeri contains an isoenzyme of MtaA designated MtbA. The isoenzyme reacted with MtaC with only 2.5 % of the activity of MtaA.

https://febs.onlinelibrary.wiley.com/doi/epdf/10.1111/j.1432-1033.1997.t01-1-00670.x

Methanol:coenzyme M methyltransferase from Methanosarcina barkeri. Purification, properties and encoding genes of the corrinoid protein MT1.

Methanogenic bacteria comprise a specialized group of obligately anaerobic microorganisms able to reduce a limited number of substrates to CH 4 . The intermediates involved in this reduction process remain bound to a series of typical C 1 -carriers. Reducing equivalents are either obtained from the oxidation of H 2 or from oxidation of carbon substrates to CO 2 . Electron transfer reactions thus constitute the very essence of the process of methanogenesis. In recent years much progress has been made in the elucidation of the special metabolic pathways and the nature of the C 1 -carriers involved in methanogenic bacteria. The energy generated at the oxidoreduction reactions, notably at the methylreductase step, is conserved by ATP synthesis. The energy is used for cell carbon synthesis and, in catalytic amounts, for the reductive activation of some methanogenic enzymes. Before the condensing reaction resulting in the formation of acetyl-CoA takes place, 2 C 1 -units are reduced or oxidized depending on the substrate to a carbonyl and a -CH 3 group. Formation of the latter proceeds via the methanogenic route. Intermediary cell carbon synthesis starting from acetyl-CoA involves reductive carboxylations and oxidoreductions by the participation of the enzymes of the tricarboxylic acid cycle.

Electron transfer reactions in methanogens

Methanosarcina barkeri is known to contain two methyltransferase isoenzymes, here designated MtaA and MtbA, which catalyze the formation of methyl-coenzyme M from methylcobalamin and coenzyme M The genes encoding the two soluble 34-kDa proteins have been cloned and sequenced mtaA and mtbA were found to be located in different parts of the genome, each forming a monocystronic transcription unit Northern blot analysis revealed that mtaA is preferentially transcribed when M barkeri is grown on methanol and the mtbA gene when the organism is grown on H2/CO2 or trimethylamine Comparison of the deduced amino acid sequences revealed the sequences of the two isoenzymes to be 37% identical Both isoenzymes showed sequence similarity to uroporphyrinogen III decarboxylase from Escherichia coli The mtaA gene was tagged with a sequence encoding six His placed six bp before the mtaA start codon, and was functionally overexpressed in E coli 25% of the E coli protein was found to be active methyltransferase which could be, purified in two steps to apparent homogenity with a 70% yield

https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1996.00653.x

Methylcobalamin: coenzyme M methyltransferase isoenzymes MtaA and MtbA from Methanosarcina barkeri. Cloning, sequencing and differential transcription of the encoding genes, and functional overexpression of the mtaA gene in Escherichia coli.

Chris van der Lest

Papers

Reductive activation of methanol: 5-hydroxybenzimidazolylcobamide methyltransferase of Methanosarcina barkeri.