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Characterization of flagellum gene families of methanogenic archaea and localization of novel flagellum accessory proteins.

ABSTRACT: Archaeal flagella are unique motility structures, and the absence of bacterial structural motility genes in the complete genome sequences of flagellated archaeal species suggests that archaeal flagellar biogenesis is likely mediated by novel components. In this study, a conserved flagellar gene family from each of Methanococcus voltae, Methanococcus maripaludis, Methanococcus thermolithotrophicus, and Methanococcus jannaschii has been characterized. These species possess multiple flagellin genes followed immediately by eight known and supposed flagellar accessory genes, flaCDEFGHIJ. Sequence analyses identified a conserved Walker box A motif in the putative nucleotide binding proteins FlaH and FlaI that may be involved in energy production for flagellin secretion or assembly. Northern blotting studies demonstrated that all the species have abundant polycistronic mRNAs corresponding to some of the structural flagellin genes, and in some cases several flagellar accessory genes were shown to be cotranscribed with the flagellin genes. Cloned flagellar accessory genes of M. voltae were successfully overexpressed as His-tagged proteins in Escherichia coli. These recombinant flagellar accessory proteins were affinity purified and used as antigens to raise polyclonal antibodies for localization studies. Immunoblotting of fractionated M. voltae cells demonstrated that FlaC, FlaD, FlaE, FlaH, and FlaI are all present in the cell as membrane-associated proteins but are not major components of isolated flagellar filaments. Interestingly, flaD was found to encode two proteins, each translated from a separate ribosome binding site. These protein expression data indicate for the first time that the putative flagellar accessory genes of M. voltae, and likely those of other archaeal species, do encode proteins that can be detected in the cell.

SUBMITTER: Thomas NA

PROVIDER: S-EPMC95564 | biostudies-literature | 2001 Dec

REPOSITORIES: biostudies-literature

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Characterization of flagellum gene families of methanogenic archaea and localization of novel flagellum accessory proteins.

Thomas N A NA Jarrell K F KF

Journal of bacteriology 20011201 24

Archaeal flagella are unique motility structures, and the absence of bacterial structural motility genes in the complete genome sequences of flagellated archaeal species suggests that archaeal flagellar biogenesis is likely mediated by novel components. In this study, a conserved flagellar gene family from each of Methanococcus voltae, Methanococcus maripaludis, Methanococcus thermolithotrophicus, and Methanococcus jannaschii has been characterized. These species possess multiple flagellin genes ...[more]

PMID: 11717274

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Project description:BackgroundArchaea perform critical roles in the microbiome system, including utilizing hydrogen to allow for enhanced microbiome member growth and influencing overall host health. With the majority of microbiome research focusing on bacteria, the functions of archaea are largely still under investigation. Understanding methanogenic functions during the host lifetime will add to the limited knowledge on archaeal influence on gut and host health. In our study, we determined lifelong archaea dynamics, including detection and methanogenic functions, while assessing global, temporal and host distribution of our novel archaeal metagenome-assembled genomes (MAGs). We followed 7 monogastric swine throughout their life, from birth to adult (1-156 days of age), and collected feces at 22 time points. The samples underwent gDNA extraction, Illumina sequencing, bioinformatic quality and assembly processes, MAG taxonomic assignment and functional annotation. MAGs were utilized in downstream phylogenetic analysis for global, temporal and host distribution in addition to methanogenic functional potential determination.ResultsWe generated 1130 non-redundant MAGs, representing 588 unique taxa at the species level, with 8 classified as methanogenic archaea. The taxonomic classifications were as follows: orders Methanomassiliicoccales (5) and Methanobacteriales (3); genera UBA71 (3), Methanomethylophilus (1), MX-02 (1), and Methanobrevibacter (3). We recovered the first US swine Methanobrevibacter UBA71 sp006954425 and Methanobrevibacter gottschalkii MAGs. The Methanobacteriales MAGs were identified primarily during the young, preweaned host whereas Methanomassiliicoccales primarily in the adult host. Moreover, we identified our methanogens in metagenomic sequences from Chinese swine, US adult humans, Mexican adult humans, Swedish adult humans, and paleontological humans, indicating that methanogens span different hosts, geography and time. We determined complete metabolic pathways for all three methanogenic pathways: hydrogenotrophic, methylotrophic, and acetoclastic. This study provided the first evidence of acetoclastic methanogenesis in archaea of monogastric hosts which indicated a previously unknown capability for acetate utilization in methanogenesis for monogastric methanogens. Overall, we hypothesized that the age-associated detection patterns were due to differential substrate availability via the host diet and microbial metabolism, and that these methanogenic functions are likely crucial to methanogens across hosts. This study provided a comprehensive, genome-centric investigation of monogastric-associated methanogens which will further improve our understanding of microbiome development and functions.

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Dataset Information

Characterization of flagellum gene families of methanogenic archaea and localization of novel flagellum accessory proteins.

Publications

Characterization of flagellum gene families of methanogenic archaea and localization of novel flagellum accessory proteins.

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