Reduction of the Pentavalent Arsenical Dimethylarsinic Acid and the GSTO1 Substrate S-(4-Nitrophenacyl)glutathione by Rat Liver Cytosol: Analyzing the Role of GSTO1 in Arsenic Reduction

Dimethylarsinic acid (DMAsv) is the major urinary metabolite of inorganic arsenic. The relatively atoxic DMAsv is reduced in the body to the much more toxic and thiol-reactive dimethylarsinous acid (DMAsIII). Glutathione S-transferase omega 1 (GSTO1) can catalyze this toxification step; however, its role in the reduction of DMAsv in vivo or by tissue extracts is unclear. Therefore, we assessed the role of GSTO1 in the reduction of DMAsv to DMAsIII by rat liver cytosol. The experiments revealed that glutathione (GSH) supported the cytosolic DMAsv reduction specifically and that GSH analogues and GSH conjugates, such as S-alkylglutathiones and S-(4-nitrophenacyl)glutathione (4-NPG; a GSTO1 specific substrate), inhibited the formation of DMAsIII. Observations in line with the view that GSTO1 catalyzes the cytosolic reduction of DMAsv include (i) findings pointing to the presence of a GSH-binding site on the DMAsv-reducing cytosolic enzyme, (ii) identical responsiveness of the DMAsv- and 4-NPG-reducing activities in rat liver cytosol to the GSTO1 specific inhibitors KTS3 and chloromethylfluorescein diacetate, and (iii) perfect coelution of the two activities during affinity and anion exchange chromatography of cytosolic proteins. Other observations appear ambiguous as to the role of GSTO1 in the cytosolic reduction of DMAsv. These include the different sensitivities of the DMAsv-reducing and GSTO1 activities to aurothioglucose, trivalent antimony, and zinc ions, as well as the preserved GSTO1 activity in cytosols whose DMAsv-reducing activity was lost due to spontaneous thiol oxidation. These disparate findings may be reconciled by assuming that GSTO1 catalyzes the reduction of both DMAsv and 4-NPG in rat liver cytosol; however, the enzyme employs different sites and/or mechanisms when reducing these substrates.