A New Approach for Dynamics of Enzyme-Catalyzed Glutathione Conjugation by Electrospray Quadrupole/ Time-of-Flight Mass Spectrometry

✍ Scribed by Masaki Ishigai; James I. Langridge; Robert S. Bordoli

• Publisher: Elsevier Science
• Year: 2001
• Tongue: English
• Weight: 151 KB
• Volume: 298
• Category: Article
• ISSN: 0003-2697
• DOI: 10.1006/abio.2001.5339

✦ Synopsis

The dynamics of enzyme-catalyzed glutathione conjugation was studied by electrospray quadrupole/ time-of-flight (Q-TOF) mass spectrometry with a nanospray interface. After incubation of human glutathione S-transferase A1-1 (GT) with glutathione (GSH) and an electrophilic substrate, electrospray indicated the presence of enzyme/product adducts such as

and [2GT ؉ 2 products] as well as [2GT] and [2GT ؉ GSH].

The relative abundance of GT/product adduct ions increased with incubation time. The wide m/z range of detection (m/z 300 -5000) allowed the observation of product, suggested to be released from enzyme/product adducts, in the same mass spectrum. The noncovalent complexes of GT/product were completely replaced by GT/inhibitor complexes following the addition of GT inhibitor to the incubation mixture. Furthermore, a collision-activated decomposition analysis of these ion species provided us with useful information to interpret or identify ion species. The results suggest that electrospray Q-TOF mass spectrometry is a powerful approach for studying the dynamics of the enzyme reaction as well as the structure of enzyme complexes at high sensitivity.