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Tau Protein pp 237–248Cite as

Production of O-GlcNAc Modified Recombinant Tau in E. coli and Detection of Ser400 O-GlcNAc Tau In Vivo

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Part of the book series: Methods in Molecular Biology ((MIMB,volume 1523))

Abstract

Assembly of the microtubule-associated protein tau (tau) into paired helical filaments that ultimately give rise to neurofibrillary tangles (NFTs) makes up one half of the two hallmark pathologies of Alzheimer’s disease (AD). Tau has been shown to be modified with O-linked N-acetylglucosamine residues (O-GlcNAc), which is the modification of serine and threonine residues of nucleocytoplasmic proteins with N-acetyl-d-glucosamine (GlcNAc) moieties. Increasing O-GlcNAc in mouse models of tauopathy has been shown to hinder the progression of symptoms in these mice and impair the aggregation of tau into NFTs. In order to study how O-GlcNAc on tau may contribute to the protective effects observed in tauopathy mouse models, it is beneficial to study O-GlcNAc modified tau in vitro. Here we describe a method for producing, purifying and enriching recombinant tau that is O-GlcNAc modified. These methods have enabled the identification of O-GlcNAc modification sites on tau including Ser400. We further describe the detection of Ser400 O-GlcNAc on tau from brain lysates.

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Acknowledgments

S.A.Y. would like to thank the National Science and Engineering Research Council of Canada (NSERC) and the Alzheimer Society of Canada for providing doctoral fellowship support while these methods were being established. D.J.V. would like to thank the Alzheimer Society of Canada and the Canadian Institutes for Health Research (MOP-123341) for support of this research and the Canada Research Chairs program for support as a Tier I Canada Research Chair in Chemical Biology.

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Authors and Affiliations

  1. Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Dr, Burnaby, BC, Canada, V5A 1S6

    Scott A. Yuzwa & David J. Vocadlo

  2. Neurosciences and Mental Health program, The Hospital for Sick Children, Toronto, ON, Canada, M5G 0A4

    Scott A. Yuzwa

  3. Department of Chemistry, Simon Fraser University, 8888 University Dr, Burnaby, BC, Canada, V5A 1S6

    David J. Vocadlo

Authors
  1. Scott A. Yuzwa
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  2. David J. Vocadlo
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Corresponding authors

Correspondence to Scott A. Yuzwa or David J. Vocadlo .

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Editors and Affiliations

  1. UMR CNRS 8576, Villeneuve d’Ascq, France

    Caroline Smet-Nocca

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Yuzwa, S.A., Vocadlo, D.J. (2017). Production of O-GlcNAc Modified Recombinant Tau in E. coli and Detection of Ser400 O-GlcNAc Tau In Vivo. In: Smet-Nocca, C. (eds) Tau Protein. Methods in Molecular Biology, vol 1523. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6598-4_13

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  • DOI: https://doi.org/10.1007/978-1-4939-6598-4_13

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6596-0

  • Online ISBN: 978-1-4939-6598-4

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