Synapsin-1 and tau reciprocal O-GlcNAcylation and phosphorylation sites in mouse brain synaptosomes.

Synapsin-1 and tau reciprocal O-GlcNAcylation and phosphorylation sites in mouse brain synaptosomes. Exp Mol Med. 2013;45:e29 Authors: Kang MJ, Kim C, Jeong H, Cho BK, Ryou AL, Hwang D, Mook-Jung I, Yi EC Abstract O-linked N-acetylglucosamine (O-GlcNAc) represents a key regulatory post-translational modification (PTM) that is reversible and often reciprocal with phosphorylation of serine and threonine at the same or nearby residues. Although recent technical advances in O-GlcNAc site-mapping methods combined with mass spectrometry (MS) techniques have facilitated study of the fundamental roles of O-GlcNAcylation in cellular processes, an efficient technique for examining the dynamic, reciprocal relationships between O-GlcNAcylation and phosphorylation is needed to provide greater insights into the regulatory functions of O-GlcNAcylation. Here, we describe a strategy for selectively identifying both O-GlcNAc- and phospho-modified sites. This strategy involves metal affinity separation of O-GlcNAcylated and phosphorylated peptides, β-elimination of O-GlcNAcyl or phosphoryl functional groups from the separated peptides followed by dithiothreitol (DTT) conjugation (BEMAD), affinity purification of DTT-conjugated peptides using thiol affinity chromatography, and identification of formerly O-GlcNAcylated or phosphorylated peptides by MS. The combined metal affinity separation and BEMAD approach allows selective enrichment of O-GlcNAcylated peptides...
Source: exp Mol Med - Category: Molecular Biology Authors: Tags: Exp Mol Med Source Type: research