Dataset Information

Genome-scale analysis of gene function in the hydrogenotrophic methanogenic archaeon Methanococcus maripaludis.

ABSTRACT: A comprehensive whole-genome analysis of gene function by transposon mutagenesis and deep sequencing methodology has been implemented successfully in a representative of the Archaea domain. Libraries of transposon mutants were generated for the hydrogenotrophic, methanogenic archaeon Methanococcus maripaludis S2 using a derivative of the Tn5 transposon. About 89,000 unique insertions were mapped to the genome, which allowed for the classification of 526 genes or about 30% of the genome as possibly essential or strongly advantageous for growth in rich medium. Many of these genes were homologous to eukaryotic genes that encode fundamental processes in replication, transcription, and translation, providing direct evidence for their importance in Archaea. Some genes classified as possibly essential were unique to the archaeal or methanococcal lineages, such as that encoding DNA polymerase PolD. In contrast, the archaeal homolog to the gene encoding DNA polymerase B was not essential for growth, a conclusion confirmed by construction of an independent deletion mutation. Thus PolD, and not PolB, likely plays a fundamental role in DNA replication in methanococci. Similarly, 121 hypothetical ORFs were classified as possibly essential and likely play fundamental roles in methanococcal information processing or metabolism that are not established outside this group of prokaryotes.

SUBMITTER: Sarmiento F

PROVIDER: S-EPMC3607031 | biostudies-literature | 2013 Mar

REPOSITORIES: biostudies-literature

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Genome-scale analysis of gene function in the hydrogenotrophic methanogenic archaeon Methanococcus maripaludis.

Sarmiento Felipe F Mrázek Jan J Whitman William B WB

Proceedings of the National Academy of Sciences of the United States of America 20130304 12

A comprehensive whole-genome analysis of gene function by transposon mutagenesis and deep sequencing methodology has been implemented successfully in a representative of the Archaea domain. Libraries of transposon mutants were generated for the hydrogenotrophic, methanogenic archaeon Methanococcus maripaludis S2 using a derivative of the Tn5 transposon. About 89,000 unique insertions were mapped to the genome, which allowed for the classification of 526 genes or about 30% of the genome as possib ...[more]

PMID: 23487778

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Project description:Hydrogenotrophic methanogenesis occurs in multiple environments, ranging from the intestinal tracts of animals to anaerobic sediments and hot springs. Energy conservation in hydrogenotrophic methanogens was long a mystery; only within the last decade was it reported that net energy conservation for growth depends on electron bifurcation. In this work, we focus on Methanococcus maripaludis, a well-studied hydrogenotrophic marine methanogen. To better understand hydrogenotrophic methanogenesis and compare it with methylotrophic methanogenesis that utilizes oxidative phosphorylation rather than electron bifurcation, we have built iMR539, a genome scale metabolic reconstruction that accounts for 539 of the 1,722 protein-coding genes of M. maripaludis strain S2. Our reconstructed metabolic network uses recent literature to not only represent the central electron bifurcation reaction but also incorporate vital biosynthesis and assimilation pathways, including unique cofactor and coenzyme syntheses. We show that our model accurately predicts experimental growth and gene knockout data, with 93% accuracy and a Matthews correlation coefficient of 0.78. Furthermore, we use our metabolic network reconstruction to probe the implications of electron bifurcation by showing its essentiality, as well as investigating the infeasibility of aceticlastic methanogenesis in the network. Additionally, we demonstrate a method of applying thermodynamic constraints to a metabolic model to quickly estimate overall free-energy changes between what comes in and out of the cell. Finally, we describe a novel reconstruction-specific computational toolbox we created to improve usability. Together, our results provide a computational network for exploring hydrogenotrophic methanogenesis and confirm the importance of electron bifurcation in this process.ImportanceUnderstanding and applying hydrogenotrophic methanogenesis is a promising avenue for developing new bioenergy technologies around methane gas. Although a significant portion of biological methane is generated through this environmentally ubiquitous pathway, existing methanogen models portray the more traditional energy conservation mechanisms that are found in other methanogens. We have constructed a genome scale metabolic network of Methanococcus maripaludis that explicitly accounts for all major reactions involved in hydrogenotrophic methanogenesis. Our reconstruction demonstrates the importance of electron bifurcation in central metabolism, providing both a window into hydrogenotrophic methanogenesis and a hypothesis-generating platform to fuel metabolic engineering efforts.

Dataset Information

Genome-scale analysis of gene function in the hydrogenotrophic methanogenic archaeon Methanococcus maripaludis.

Publications

Genome-scale analysis of gene function in the hydrogenotrophic methanogenic archaeon Methanococcus maripaludis.

Similar Datasets

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