A novel N-linked flagellar glycan from Methanococcus maripaludis

Area of Science:

• Microbiology
• Glycobiology
• Biochemistry

Background:

• Archaea utilize diverse post-translational modifications.
• Flagellar glycosylation is crucial for archaeal motility and cell surface properties.
• Methanococcus maripaludis Mm900 flagella are known to be glycosylated.

Purpose of the Study:

• To elucidate the structure of the N-linked glycan present on Methanococcus maripaludis Mm900 flagella.
• To characterize the novel monosaccharide unit within the tetrasaccharide.

Main Methods:

• Mass spectrometry (MS) analysis of tryptic peptides to identify glycopeptides.
• Purification of the glycan using pronase digestion, size-exclusion, and anion-exchange chromatography.
• Nuclear Magnetic Resonance (NMR) spectroscopy for detailed structural elucidation.

Main Results:

• Identification of tryptic glycopeptides with a glycan mass of 1036.4 Da.
• Confirmation of a tetrasaccharide structure through glycan oxonium ion fragmentation.
• Determination of the complete glycan structure: Sug-4-beta-ManNAc3NAmA6Thr-4-beta-GlcNAc3NAcA-3-beta-GalNAc-Asn.
• Characterization of 'Sug' as a novel monosaccharide: (5S)-2-acetamido-2,4-dideoxy-5-O-methyl-alpha-l-erythro-hexos-5-ulo-1,5-pyranose.
• Observed significant structural similarity to the oligosaccharide found in Methanococcus voltae.

Conclusions:

• The N-linked tetrasaccharide on M. maripaludis flagella is composed of known and a novel monosaccharide.
• The determined structure provides insights into archaeal glycan biosynthesis and evolution.
• This finding expands the understanding of post-translational modifications in archaea.