A less bias approach using chemoenzymatic tagging coupl with click chemistry has en develop for the investigation of protein O-GlcNAcylation (Thompson et al., 2018). Mass spectrometry-bas strategies (Alfaro et al., 2012; Wang et al., 2014; Wang et al., 2017; and Thompson et al., 2018) are now increasingly ing adopt to determine how specific protein O-GlcNAcylation is regulat. Recent studies have also gun to tackle the more challenging question of how the intracellular localization of specific O-GlcNAcylat proteins is chang in response to various physiological and pathological conditions. To provide practical considerations of the various approaches to study the role and regulation of protein O-GlcNAcylation, in this review, we cover several major areas.
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For approaches, we discuss the following four aspects: a) The determination of the activities of the enzymes OGT and OGA. b) The assessment of overall and specific O-GlcNAcylation levels in cells/tissues using antibody or click chemistry methods. c) The determination and quantification of protein O-GlcNAc modification by mass spectrometry (MS). d) The determination of intracellular localization and function of O-GlcNAcylat proteins in vitro and in vivo. For studies in humans Finland Email List and animal models that sh light on biological function of protein O-GlcNAcylation, we discuss a) observations in humans, b) observations in genetically modifi mouse models, and c) observations using potent pharmacological agents. We conclude with research questions and future directions in O-GlcNAc biology and its role in health and disease.
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Approaches for Assessing Enzymatic Activities of OGT and OGA and for Assessing the Global and Specific Protein O-GlcNAcylation To understand the functions of protein O-GlcNAcylation, it is important to able to measure the two enzymes that add and remove O-GlcNAc from proteins and the extent of overall protein O-GlcNAcylation in a given tissue or underdefin conditions. The following four sections provide an overview of these approaches. Furthermore, the development of TH Lists methods to assess the location of specific O-GlcNAcylat proteins and the function of specific modifications in vitro and in vivo will also highlight.
