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Issue 14, 2017
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Chemoselective ligation reaction of N-acetylglucosamine (NAG) with hydrazide functional probes to determine galactosyltransferase activity by MALDI mass spectrometry

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Abstract

Quantification of β-1,4-galactosyltransferase (β-1,4-GT) activity is of considerable significance in the diagnosis of various cancers including lung and ovarian cancer. We report here the use of synthetic β-N-acetylglucosamine (NAG) ligands that contain hydrazide functional groups to determine galactosyltransferase activity by mass spectrometry. With hydrazide-linked β-D-NAG as the acceptor, the activity of β-1,4-GT is quantified by matrix-assisted laser desorption/ionization (MALDI)-time-of-flight (TOF) mass spectrometry (MS) with high efficiency. Using the disulfide moiety in a 3,3′-dithiodipropionic acid dihydrazide (DTP)-linked β-D-NAG probe, Au nanoparticles (AuNPs) are employed for enriching DTP-linked β-D-NAG after enzymatic reaction, and the ligand-bound AuNPs are subsequently deposited on a MALDI plate for analysis. In addition, we have demonstrated that a perfluorocarbon (PF) labeled β-NAG-ligand can be useful for surface-based enzymatic reaction with a perfluorooctadecanethiol (PFDT)-covered gold surface. Using the ratiometric method, the conversion rate of β-1,4-GT is determined to be 0.83 ± 0.03, which shows a high level of activity. This is the first work that uses hydrazide-linked β-D-NAG for activity analysis, providing a new surface-based MS approach to determine enzyme activity in a potentially high-throughput manner.

Graphical abstract: Chemoselective ligation reaction of N-acetylglucosamine (NAG) with hydrazide functional probes to determine galactosyltransferase activity by MALDI mass spectrometry

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Publication details

The article was received on 10 Mar 2017, accepted on 14 May 2017 and first published on 17 May 2017


Article type: Paper
DOI: 10.1039/C7AN00428A
Citation: Analyst, 2017,142, 2654-2662
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    Chemoselective ligation reaction of N-acetylglucosamine (NAG) with hydrazide functional probes to determine galactosyltransferase activity by MALDI mass spectrometry

    H. Yang and Q. Cheng, Analyst, 2017, 142, 2654
    DOI: 10.1039/C7AN00428A

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