Identification of genes involved in the assembly and attachment of a novel flagellin N-linked tetrasaccharide important for motility in the archaeon Methanococcus maripaludis

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DOIResolve DOI: http://doi.org/10.1111/j.1365-2958.2009.06671.x
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TypeArticle
Journal titleMolecular Microbiology
Volume72
Issue3
Pages633644; # of pages: 12
AbstractRecently, the flagellin proteins of Methanococcus maripaludis were found to harbour an N-linked tetrasaccharide composed of N-acetylgalactosamine, di-acetylated glucuronic acid, an acetylated and acetamidino-modified mannuronic acid linked to threonine, and a novel terminal sugar [(5S)-2- acetamido-2,4-dideoxy-5-O-methyl-α-L-erythro-hexos- 5-ulo-1,5-pyranose]. To identify genes involved in the assembly and attachment of this glycan, in-frame deletions were constructed in putative glycan assembly genes. Successful deletion of genes encoding three glycosyltransferases and an oligosaccharyltransferase (Stt3p homologue) resulted in flagellins of decreased molecular masses as evidenced by immunoblotting, indicating partial or completely absent glycan structures. Deletion of the oligosaccharyltransferase or the glycosyltransferase responsible for the transfer of the second sugar in the chain resulted in flagellins that were not assembled into flagella filaments, as evidenced by electron microscopy. Deletions of the glycosyltransferases responsible for the addition of the third and terminal sugars in the glycan were confirmed by mass spectrometry analysis of purified flagellins from these mutants. Although flagellated, these mutants had decreased motility as evidenced by semi-swarm plate analysis with the presence of each additional sugar improving movement capabilities.
Publication date
LanguageEnglish
AffiliationNational Research Council Canada (NRC-CNRC); NRC Institute for Biological Sciences
Peer reviewedYes
NPARC number15460078
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Record identifier1f9658f3-ea92-44fe-86ba-47c542aa96cf
Record created2010-08-16
Record modified2016-05-09
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