Complementation of an aglB Mutant of Methanococcus maripaludis with Heterologous Oligosaccharyltransferases

PLoS One. 2016 Dec 1;11(12):e0167611. doi: 10.1371/journal.pone.0167611. eCollection 2016.


The oligosaccharyltransferase is the signature enzyme for N-linked glycosylation in all domains of life. In Archaea, this enzyme termed AglB, is responsible for transferring lipid carrier-linked glycans to select asparagine residues in a variety of target proteins including archaellins, S-layer proteins and pilins. This study investigated the ability of a variety of AglBs to compensate for the oligosaccharyltransferase activity in Methanococcus maripaludis deleted for aglB, using archaellin FlaB2 as the reporter protein since all archaellins in Mc. maripaludis are modified at multiple sites by an N-linked tetrasaccharide and this modification is required for archaellation. In the Mc. maripaludis ΔaglB strain FlaB2 runs as at a smaller apparent molecular weight in western blots and is nonarchaellated. We demonstrate that AglBs from Methanococcus voltae and Methanothermococcus thermolithotrophicus functionally replaced the oligosaccharyltransferase activity missing in the Mc. maripaludis ΔaglB strain, both returning the apparent molecular weight of FlaB2 to wildtype size and restoring archaellation. This demonstrates that AglB from Mc. voltae has a relaxed specificity for the linking sugar of the transferred glycan since while the N-linked glycan present in Mc. voltae is similar to that of Mc. maripaludis, the Mc. voltae glycan uses N-acetylglucosamine as the linking sugar. In Mc. maripaludis that role is held by N-acetylgalactosamine. This study also identifies aglB from Mtc. thermolithotrophicus for the first time by its activity. Attempts to use AglB from Methanocaldococcus jannaschii, Haloferax volcanii or Sulfolobus acidocaldarius to functionally replace the oligosaccharyltransferase activity missing in the Mc. maripaludis ΔaglB strain were unsuccessful.

MeSH terms

  • Acetylgalactosamine / metabolism*
  • Alanine / metabolism
  • Amino Acid Sequence / genetics
  • Carbohydrate Conformation
  • Extremophiles / genetics
  • Extremophiles / metabolism
  • Fimbriae Proteins / genetics
  • Fimbriae Proteins / metabolism
  • Glycosylation
  • Hexosyltransferases / genetics*
  • Lipids / genetics
  • Membrane Proteins / genetics*
  • Methanococcus / enzymology*
  • Methanococcus / genetics
  • Mutant Proteins / genetics*
  • Mutant Proteins / metabolism
  • Oligosaccharides / metabolism
  • Polysaccharides / genetics
  • Polysaccharides / metabolism


  • Lipids
  • Membrane Proteins
  • Mutant Proteins
  • Oligosaccharides
  • Polysaccharides
  • Fimbriae Proteins
  • Hexosyltransferases
  • dolichyl-diphosphooligosaccharide - protein glycotransferase
  • Acetylgalactosamine
  • Alanine

Grant support

Funding for the project was supplied by two separate grants from Natural Sciences and Engineering Research Council of Canada (NSERC), one to KFJ and the other to CMK. NSERC website The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.