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Glycoconjugate Journal

, Volume 35, Issue 6, pp 525–535 | Cite as

Identification and characterization of the 4-epimerase AglW from the archaeon Methanococcus maripaludis

  • Sulav Sharma
  • Yan Ding
  • Ken F. Jarrell
  • Inka Brockhausen
Original Article
  • 82 Downloads

Abstract

Archaea are ubiquitous single-cell microorganisms that have often adapted to harsh conditions and play important roles in biogeochemical cycles with potential applications in biotechnology. Methanococcus maripaludis, a methane-producing archaeon, is motile through multiple archaella on its cell surface. The major structural proteins (archaellins) of the archaellum are glycoproteins, modified with N-linked tetrasaccharides that are essential for the proper assembly and function of archaella. The aglW gene, encoding the putative 4-epimerase AglW, plays a key role in the synthesis of the tetrasaccharide. The goal of our work was to biochemically demonstrate the 4-epimerase activity of AglW, and to develop assays to determine its substrate specificity and properties. We carried out assays using UDP-Galactose, UDP-Glucose, UDP-N-acetylglucosamine, UDP-N-acetylgalactosamine and N-acetylglucosamine/N-acetylgalactosamine-diphosphate – lipid as substrates, coupled with specific glycosyltransferases. We showed that AglW has a broad specificity towards UDP-sugars and that Tyr151 within a conserved YxxxK sequon is essential for the 4-epimerase function of AglW. The glycosyltransferase-coupled assays are generally useful for the identification and specificity studies of novel 4-epimerases.

Keywords

Archaea N-glycosylation 4-epimerase Coupled assays Glycosyltransferases Specificity 

Notes

Acknowledgements

This research was supported by Discovery Grants from the Natural Sciences and Engineering Research Council of Canada (NSERC) to I.B and to K.F.J.

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Supplementary material

10719_2018_9845_MOESM1_ESM.pdf (636 kb)
ESM 1 (PDF 636 kb)

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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  1. 1.Department of Biomedical and Molecular SciencesQueen’s UniversityKingstonCanada

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