title: Production of O-GlcNAc Modified Recombinant Tau in E. coli and Detection of Ser400 O-GlcNAc Tau In Vivo.
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.
authors.affiliation: Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Dr, Burnaby, BC, Canada, V5A 1S6. syuzwa@sfu.ca.|Neurosciences and Mental Health program, The Hospital for Sick Children, Toronto, ON, Canada, M5G 0A4. syuzwa@sfu.ca.
authors.forename: Scott A
authors.initials: SA
authors.lastname: Yuzwa
authors.affiliation: Department of Molecular Biology and Biochemistry, Simon Fraser University, 8888 University Dr, Burnaby, BC, Canada, V5A 1S6. dvocadlo@sfu.ca.|Department of Chemistry, Simon Fraser University, 8888 University Dr, Burnaby, BC, Canada, V5A 1S6. dvocadlo@sfu.ca.
authors.forename: David J
authors.initials: DJ
authors.lastname: Vocadlo
mesh_terms: D000117:Acetylglucosamine / Q000737:chemistry* / Q000378:metabolism*; D000818:Animals; D001921:Brain / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006801:Humans; D066298:In Vitro Techniques; D051379:Mice; D011499:Protein Processing, Post-Translational; D051381:Rats; D017207:Rats, Sprague-Dawley; D024801:Tauopathies / Q000235:genetics / Q000378:metabolism; D016875:tau Proteins / Q000737:chemistry* / Q000378:metabolism*
chemical_list: D016875:tau Proteins; D000117:Acetylglucosamine
keywords: Microtubule-associated protein tau; O-GlcNAc; Purification; Ser400 O-GlcNAc tau; Tau