Next‐Generation Metabolic Glycosylation Reporters Enable Detection of Protein O−GlcNAcylation in Living Cells without S‐Glyco Modification

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2024
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Deutsche Forschungsgemeinschaft (DFG): SFB 969, project B05
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Angewandte Chemie International Edition. Wiley. 2024, 63(20), e202320247. ISSN 1433-7851. eISSN 1521-3773. Verfügbar unter: doi: 10.1002/anie.202320247
Zusammenfassung

Protein O−GlcNAcylation is a ubiquitous posttranslational modification of cytosolic and nuclear proteins involved in numerous fundamental regulation processes. Investigation of O−GlcNAcylation by metabolic glycoengineering (MGE) has been carried out for two decades with peracetylated N ‐acetylglucosamine (GlcNAc) and N ‐acetylgalactosamine derivatives modified with varying reporter groups. Recently, it has been shown that these derivatives can result in non‐specific protein labeling termed S ‐glyco modification. Here, we report norbornene‐modified GlcNAc derivatives with a protected phosphate at the anomeric position and their application in MGE. These derivatives overcome two limitations of previously used O−GlcNAc reporters. They do not lead to detectable S ‐glyco modification, and they efficiently react in the inverse‐electron‐demand Diels–Alder (IEDDA) reaction, which can be carried out even within living cells. Using a derivative with an S ‐acetyl‐2‐thioethyl‐protected phosphate, we demonstrate the protein‐specific detection of O−GlcNAcylation of several proteins and the protein‐specific imaging of O−GlcNAcylation inside living cells by Förster resonance energy transfer (FRET) visualized by confocal fluorescence lifetime imaging microscopy (FLIM).

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540 Chemie
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bioorthogonal chemistry, carbohydrates, FRET, glycoproteins, metabolic engineering
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ISO 690KUFLEITNER, Markus, Lisa Maria HAIBER, Shuang LI, Harsha SURENDRAN, Thomas U. MAYER, Andreas ZUMBUSCH, Valentin WITTMANN, 2024. Next‐Generation Metabolic Glycosylation Reporters Enable Detection of Protein O−GlcNAcylation in Living Cells without S‐Glyco Modification. In: Angewandte Chemie International Edition. Wiley. 2024, 63(20), e202320247. ISSN 1433-7851. eISSN 1521-3773. Verfügbar unter: doi: 10.1002/anie.202320247
BibTex
@article{Kufleitner2024-05-13NextG-69845,
  year={2024},
  doi={10.1002/anie.202320247},
  title={Next‐Generation Metabolic Glycosylation Reporters Enable Detection of Protein O−GlcNAcylation in Living Cells without <i>S</i>‐Glyco Modification},
  number={20},
  volume={63},
  issn={1433-7851},
  journal={Angewandte Chemie International Edition},
  author={Kufleitner, Markus and Haiber, Lisa Maria and Li, Shuang and Surendran, Harsha and Mayer, Thomas U. and Zumbusch, Andreas and Wittmann, Valentin},
  note={Article Number: e202320247}
}
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