Dataset Information

New mode of methanogenesis observed in the coal degrading methanogen Methermicoccus shengliensis

ABSTRACT: Methane-generating Archaea drive the final step in anaerobic organic compound mineralization and dictate the carbon flow of Earth’s diverse anaerobic ecosystems. Although such Archaea were presumed to be restricted to life on simple compounds like H2, acetate or methanol, an archaeon, Methermicoccus shengliensis, was recently found to convert methoxylated aromatic compounds to methane. Methoxylated aromatic compounds as component of lignin and coal are present in most subsurface sediments. Despite the significance and novelty of this outstanding archaeon its metabolism has not yet been explored. In this study, transcriptomics and proteomics reveal that M. shengliensis uses a demethoxylation system that is more related to that from acetogenic bacteria than to the methyl transferase system used for methylotrophic methanogenesis. It activates methoxy-groups using tetrahydromethanopterin as the carrier, a mechanism distinct from conventional methanogenic methyl-transfer systems dependent on Coenzyme M.

INSTRUMENT(S):

ORGANISM(S): Methermicoccus Shengliensis Dsm 18856

SUBMITTER: Julia Kurth

LAB HEAD: Jeppe Lund Nielsen

PROVIDER: PXD018934 | Pride | 2021-06-20

REPOSITORIES: Pride

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Methanogenic archaea use a bacteria-like methyltransferase system to demethoxylate aromatic compounds.

Kurth Julia M JM Nobu Masaru K MK Tamaki Hideyuki H de Jonge Nadieh N Berger Stefanie S Jetten Mike S M MSM Yamamoto Kyosuke K Mayumi Daisuke D Sakata Susumu S Bai Liping L Cheng Lei L Nielsen Jeppe Lund JL Kamagata Yoichi Y Wagner Tristan T Welte Cornelia U CU

The ISME journal 20210618 12

Methane-generating archaea drive the final step in anaerobic organic compound mineralization and dictate the carbon flow of Earth's diverse anoxic ecosystems in the absence of inorganic electron acceptors. Although such Archaea were presumed to be restricted to life on simple compounds like hydrogen (H<sub>2</sub>), acetate or methanol, an archaeon, Methermicoccus shengliensis, was recently found to convert methoxylated aromatic compounds to methane. Methoxylated aromatic compounds are important ...[more]

PMID: 34145392

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Project description:Our goal is to convert methane efficiently into liquid fuels that may be more readily transported. Since aerobic oxidation of methane is less efficient, we focused on anaerobic processes to capture methane, which are accomplished by anaerobic methanotrophic archaea (ANME) in consortia. However, no pure culture capable of oxidizing and growing on methane anaerobically has been isolated. In this study, Methanosarcina acetivorans, an archaeal methanogen, was metabolically engineered to take up methane, rather than to generate it. To capture methane, we cloned the DNA coding for the enzyme methyl-coenzyme M reductase (Mcr) from an unculturable archaeal organism from a Black Sea mat into M. acetivorans to effectively run methanogenesis in reverse. The engineered strain produces primarily acetate, and our results demonstrate that pure cultures can grow anaerobically on methane. Differential gene analysis of two growth conditions (three biological replicates each) was performed: (i) M. acetivorans/pES1-MATmcr3 grown on methane and (ii) M. acetivorans/pES1-MATmcr3 grown on methanol. All starter cultures (200 mL) were grown on methanol for 5 days, and harvested by centrifugation. Cell pellets were washed three times with HS medium, and resuspended using 5 mL HS medium, 2 Âµg/mL puromycin, and 0.1 mM FeCl3. For condition (i), methane was filled into the headspace of the cultures. For condition (ii), 150 mM methanol was added. All cultures were incubated at 37C for 5 days, followed by rapid centrifugation in the presence of 50 ÂµL RNAlater solution (Ambion, Austin, TX) per mL of culture. Total RNA was isolated using RNeasy Mini kit (Qiagen, Valencia, CA) were then digested with terminator 5â-phosphate-dependent exonuclease (Epicentre, Madison, WI) to partially remove ribosomal RNA. Digested RNA were cleaned up using AgenCourt RNAClean XP beads (AgenCourt Bioscience, Beverly, MA) and used for cDNA library construction using the TruSeq Stranded mRNA Library kit (Illumina). Pooled and barcoded cDNA library was then sequenced on a HiSeq sequencing platform (Illumina). Obtained reads were mapped to the reference genome of M. acetivorans (Genbank accession NC_003552.1) using STAR. The mapped reads were assembled using Cufflink v2.2.1 to identify potential novel transcripts. Assembled, unannotated novel transcripts for all the strains were combined with the list of known genes. Differential expression of genes and potential novel transcripts were determined using Cuffdiff at a significance cutoff at q < 0.07 with a false discovery rate of 0.05. Expression levels of gene transcripts are expressed as fragments per kilobase of transcript per million mapped fragments (FPKM), and expression changes are determined by the ratio of FPKM of culture replicates grown on methane to FPKM of culture replicates grown on methanol.

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