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Detecting and Imaging O-GlcNAc Sites Using Glycosyltransferases: A Systematic Approach to Study O-GlcNAc

24 Pages Posted: 9 Jun 2018 Publication Status: Published

See all articles by Zhengliang L. Wu

Zhengliang L. Wu

R&D Systems, Inc. - Bio-Techne Ltd.

Timothy J. Tatge

R&D Systems, Inc. - Bio-Techne Ltd.

Alex Grill

R&D Systems, Inc. - Bio-Techne Ltd.

Yonglong Zhou

R&D Systems, Inc. - Bio-Techne Ltd.

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Abstract

O-GlcNAcylation is a reversible serine/threonine glycosylation for regulating protein activity and availability inside of cells. In a given protein, O-GlcNAcylated and unoccupied O-GlcNAc sites are referred as closed and open sites, respectively. The balance between open and closed sites is believed to be dynamically regulated. In this report, closed sites are detected using in vitro incorporation of GalNAz by B3GALNT2, and open sites are detected by in vitro incorporation of GlcNAz by OGT, via click chemistry. For assessing total O-GlcNAc sites, a sample is OGlcNAcylated in vitro by OGT before detecting by B3GALNT2. The methods are demonstrated on purified protein, cellular extracts, and whole cells. Through O-GlcNAc imaging, the modification degree of O-GlcNAc in nuclei of CHO cells was estimated. The detection and imaging of both open and closed O-GlcNAc sites provide a systematic approach to study this important post-translational modification.

Suggested Citation

Wu, Zhengliang L. and Tatge, Timothy J. and Grill, Alex and Zhou, Yonglong, Detecting and Imaging O-GlcNAc Sites Using Glycosyltransferases: A Systematic Approach to Study O-GlcNAc (2018). Available at SSRN: https://ssrn.com/abstract=3188488 or http://dx.doi.org/10.2139/ssrn.3188488
This version of the paper has not been formally peer reviewed.

Zhengliang L. Wu (Contact Author)

R&D Systems, Inc. - Bio-Techne Ltd. ( email )

19 Barton Lane
Abingdon Science Park
Abingdon, OX14 3NB
United Kingdom

Timothy J. Tatge

R&D Systems, Inc. - Bio-Techne Ltd.

19 Barton Lane
Abingdon Science Park
Abingdon, OX14 3NB
United Kingdom

Alex Grill

R&D Systems, Inc. - Bio-Techne Ltd.

19 Barton Lane
Abingdon Science Park
Abingdon, OX14 3NB
United Kingdom

Yonglong Zhou

R&D Systems, Inc. - Bio-Techne Ltd.

19 Barton Lane
Abingdon Science Park
Abingdon, OX14 3NB
United Kingdom

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