Simple fluorescent reagents for monitoring disulfide reductase and S-nitroso reductase activities in vitro and in live cells in culture

Abstract

This study describes the theoretical basis and the methods for the facile synthesis and characterization of four fluorogenic probes, N-amido-O-aminobenzoyl-S-nitrosoglutathione (AOASNOG), N-thioamido-fluoresceinyl-S-nitroso-glutathione (TFSNOG), N,N-di(thioamido-fluoresceinyl)-cystine (DTFCys₂) and N,N-di(thioamido-fluoresceinyl)-homocystine (DTFHCys₂). In addition, the study describes the methodology for the application of these reagents for measuring and imaging of free thiols on cell surfaces as well as their use as pseudo substrates for the thiol reductase and S-nitrosothioldenitrosylase activities of protein disulfide isomerase (PDI) and S-nitrosothiol reductase activity of S-nitrosoglutathione reductase (GSNOR) in vitro and on live cells in culture.

Keywords: Fluorogenic thiol redox probes; Protein disulfide isomerase; S-Nitroglutathione reductase.