Chemoselective ligation reaction of N-acetylglucosamine (NAG) with hydrazide functional probes to determine galactosyltransferase activity by MALDI mass spectrometry. Issue 14 (15th June 2017)

• Record Type: Journal Article
• Authors: Yang, Hyojik; Cheng, Quan
• Is Part Of: Analyst. Volume 142:Issue 14(2017)
• Page Start: 2654
• Page End: 2662
• Publication Date: 2017-06-15
• Subjects: Chemistry, Analytic -- Periodicals; 543
• DOI: 10.1039/c7an00428a
• Languages: English
• ISSNs: 0003-2654

Abstract

Abstract : A perfluorocarbon-modified gold surface is used to immobilize PF-β-NAG and allows quantification of β-GT enzymatic activity with MALDI-TOF/MS. 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.