Project description:Population dynamics of methanogenic genera was investigated in pilot anaerobic digesters. Cattle manure and two-phase olive mill wastes were codigested at a 3:1 ratio in two reactors operated at 37 ï¾°C and 55 ï¾°C. Other two reactors were run with either residue at 37 ï¾°C. Sludge DNA extracted from samples taken from all four reactors on days 4, 14 and 28 of digestion was used for hybridisation with the AnaeroChip, an oligonucleotide microarray targeting those groups of methanogenic archaea that are commonly found under mesophilic and thermophilic conditions (Franke-Whittle et al. 2009, in press, doi:10.1016/j.mimet.2009.09.017).
Project description:Benedict2011 - Genome-scale metoblic network
of Methanosarcina acetivorans (iMB745)
This model is described in the article:
Genome-scale metabolic
reconstruction and hypothesis testing in the methanogenic
archaeon Methanosarcina acetivorans C2A.
Benedict MN, Gonnerman MC, Metcalf
WW, Price ND.
J. Bacteriol. 2012 Feb; 194(4):
855-865
Abstract:
Methanosarcina acetivorans strain C2A is a marine
methanogenic archaeon notable for its substrate utilization,
genetic tractability, and novel energy conservation mechanisms.
To help probe the phenotypic implications of this organism's
unique metabolism, we have constructed and manually curated a
genome-scale metabolic model of M. acetivorans, iMB745, which
accounts for 745 of the 4,540 predicted protein-coding genes
(16%) in the M. acetivorans genome. The reconstruction effort
has identified key knowledge gaps and differences in peripheral
and central metabolism between methanogenic species. Using flux
balance analysis, the model quantitatively predicts wild-type
phenotypes and is 96% accurate in knockout lethality
predictions compared to currently available experimental data.
The model was used to probe the mechanisms and energetics of
by-product formation and growth on carbon monoxide, as well as
the nature of the reaction catalyzed by the soluble
heterodisulfide reductase HdrABC in M. acetivorans. The
genome-scale model provides quantitative and qualitative
hypotheses that can be used to help iteratively guide
additional experiments to further the state of knowledge about
methanogenesis.
This model is hosted on
BioModels Database
and identified by:
MODEL1507180040.
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To the extent possible under law, all copyright and related or
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Project description:Methanogenesis allows methanogenic archaea (methanogens) to generate cellular energy for their growth while producing methane. Hydrogenotrophic methanogens thrive on carbon dioxide and molecular hydrogen as sole carbon and energy sources. Thermophilic and hydrogenotrophic Methanothermobacter spp. have been recognized as robust biocatalysts for a circular carbon economy and are now applied in power-to-gas technology. Here, we generated the first manually curated genome-scale metabolic reconstruction for three Methanothermobacter spp.. We investigated differences in growth performance of three wild-type strains and one genetically engineered strain in two independent chemostat bioreactor experiments, first, with molecular hydrogen and carbon dioxide, and second, with sodium formate. In the first experiment, we found the highest methane production rate for Methanothermobacter thermautotrophicus ΔH, while Methanothermobacter marburgensis Marburg reached the highest biomass growth rate. Transcriptomics and proteomics data sets from these steady-state bioreactors, in combination with the implementation of a pan-model that contains combined reactions from all three microbes, allowed us to perform an interspecies comparison. While the observed differences in the growth behavior cannot be fully explained, the comparison enabled us to identify crucial differences in growth-related pathways such as formate anabolism. In the second experiment, we found stable growth with a M. thermautotrophicus ΔH plasmid-carrying strain on formate with similar performance parameters compared to wild-type Methanothermobacter thermautotrophicus Z-245. The results of the two studies demonstrate the advantages of an integrative approach combining fermentation and omics data with genome-scale modeling to reveal knowledge gaps of archaeal metabolisms and the biotechnological potential of Methanothermobacter spp. as production platform hosts.
Project description:Recent advances in the study of archaeal DNA replication have uncovered defined replication origins (oriC) and demonstrated specific binding of the Cdc6/Orc1 protein and Mcm helicase to oriC in vivo and/or in vitro. The oriC of the hyperthermophilic archaeon Pyrococcus abyssi is characterized by 13 bp repeats, AT-rich regions and an inverted repeat whose precise roles remain unclear. We report here that the 13 bp repeats are widespread in the three Pyrococcus genomes. Nevertheless, by means of chromatin immunoprecipitation coupled with hybridization on a whole genome microarray (ChIP-chip), we found that binding of P. abyssi Cdc6/Orc1 to oriC in vivo was highly specific both in exponential and stationary phases, allowing oriC to be distinguished in the 1.8 M bp genome. ChIP-chip analysis also indicated that a single 13 bp repeat is not sufficient for stable binding of Cdc6/Orc1. Purified Cdc6/Orc1 binds a DNA fragment containing the inverted repeat of oriC with a relatively low affinity, suggesting that multiple clusters of the 13 bp repeat discovered in this study contribute to the stable binding of Cdc6/Orc1 to oriC. Our ChIP-chip analysis revealed that Mcm also binds oriC only in proliferating cells, consistent with its role as a licensing factor. Finally, we found that both Cdc6/Orc1 and Mcm have one additional target site. Notably, Mcm binds constitutively to a GC-rich region containing two rRNA genes and a tRNA gene, suggesting a role of archaeal Mcm in DNA replication and/or transcription of this peculiar region. Keywords: ChIP-chip
Project description:Methanococcus maripaludis is a methanogenic archaeon. Within its genome, there are two operons for membrane associated hydrogenases, eha and ehb. To investigate the regulation of ehb on the cell, an S40 mutant was constructed in such a way that a portion of the ehb operon was replaced by pac cassette in the wild type parental strain S2 (done by Whitman's group at the University of Georgia). Four samples of each strain were grown in batch culture. Differences in transcriptional expression between S40 and S2 were measured using cDNA arrays, with flip dye experiments for each biological replicate. Keywords: mutant, archaea, carbon, hydrogenase, methanogen