Regulation of microsomal and cytosolic glutathione S-transferase activities by S-nitrosylation.
There is increasing evidence that S-nitrosylation is a mechanism for the regulation of protein function via the modification of critical sulfhydryl groups. The activity of rat liver microsomal glutathione S-transferase (GST) is increased after treatment with N-ethylmaleimide (NEM), a sulfhydryl alkylating reagent, and is also increased under conditions of oxidative stress. In the present study, preincubation of purified rat liver microsomal GST with S-nitrosoglutathione (GSNO) or the nitric oxide (NO) donor, 1,1-diethyl-2-hydroxy-2-nitrosohydrazine (DEA/NO), resulted in a 2-fold increase in enzyme activity. This increase in activity was reversed by dithiothreitol. The initial treatment of microsomal GST with either GSNO or DEA/NO was associated with an 85% loss of free sulfhydryl groups. After removal of the nitrosylating agents over a 6-hr period, approximately 50% of the enzyme was still nitrosylated, as determined by redox chemiluminescence. Furthermore, preincubation of either purified enzyme or hepatic microsomes with GSNO or DEA/NO prevented further enzyme activation by NEM, suggesting that NEM and the NO donors interact with a common population of sulfhydryl groups in the enzyme. In contrast, both NEM and NO donors partially inhibited the activity of cytosolic GST isoforms. The inhibitory activity of NEM and NO donors was much more evident when the GST pi isoform was used instead of a mixture of GST isoforms. These data suggest that there may be differential regulation of microsomal and cytosolic GST activities under conditions of nitrosative stress.
Ji
Yanbin
Y
Department of Pharmacology and Toxicology, Faculty of Health Sciences, Queen's University, Kingston, Ontario, Canada K7L 3N6.
Toader
Violeta
V
Bennett
Brian M
BM
D000818:Animals; D003600:Cytosol / Q000201:enzymology; D005982:Glutathione Transferase / Q000378:metabolism*; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D020030:Nitric Oxide Donors / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D026422:S-Nitrosoglutathione / Q000378:metabolism*
D020030:Nitric Oxide Donors; D026422:S-Nitrosoglutathione; D005982:Glutathione Transferase