Polynucleotide phosphorylase and mitochondrial ATP synthase mediate reduction of arsenate to the more toxic arsenite by forming arsenylated analogues of ADP and ATP.
We have demonstrated that phosphorolytic-arsenolytic enzymes can promote reduction of arsenate (AsV) into the more toxic arsenite (AsIII) because they convert AsV into an arsenylated product in which the arsenic is more reducible by glutathione (GSH) or other thiols to AsIII than in inorganic AsV. We have also shown that mitochondria can rapidly reduce AsV in a process requiring intact oxidative phosphorylation and intramitochondrial GSH. Thus, these organelles might reduce AsV because mitochondrial ATP synthase, using AsV instead of phosphate, arsenylates ADP to ADP-AsV, which in turn is readily reduced by GSH. To test this hypothesis, we first examined whether the RNA-cleaving enzyme polynucleotide phosphorylase (PNPase), which can split poly-adenylate (poly-A) by arsenolysis into units of AMP-AsV (a homologue of ADP-AsV), could also promote reduction of AsV to AsIII in presence of thiols. Indeed, bacterial PNPase markedly facilitated formation of AsIII when incubated with poly-A, AsV, and GSH. PNPase-mediated AsV reduction depended on arsenolysis of poly-A and presence of a thiol. PNPase can also form AMP-AsV from ADP and AsV (termed arsenolysis of ADP). In presence of GSH, this reaction also facilitated AsV reduction in proportion to AMP-AsV production. Although various thiols did not influence the arsenolytic yield of AMP-AsV, they differentially promoted the PNPase-mediated reduction of AsV, with GSH being the most effective. Circumstantial evidence indicated that AMP-AsV formed by PNPase is more reducible to AsIII by GSH than inorganic AsV. Then, we demonstrated that AsV reduction by isolated mitochondria was markedly inhibited by an ADP analogue that enters mitochondria but is not phosphorylated or arsenylated. Furthermore, inhibitors of the export of ATP or ADP-AsV from the mitochondria diminished the increment in AsV reduction caused by adding GSH externally to these organelles whose intramitochondrial GSH had been depleted. Thus, whereas PNPase promotes reduction of AsV by incorporating it into AMP-AsV, the mitochondrial ATP synthase facilitates AsV reduction by forming ADP-AsV; then GSH can easily reduce these arsenylated nucleotides to AsIII.
Németi
Balázs
B
Department of Pharmacology and Pharmacotherapy, Toxicology Section, University of Pécs, Medical School, H-7624 Pécs, Hungary.
Regonesi
Maria Elena
ME
Tortora
Paolo
P
Gregus
Zoltán
Z
D000244:Adenosine Diphosphate / Q000378:metabolism*; D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D001149:Arsenates / Q000378:metabolism*; D053501:Arsenite Transporting ATPases / Q000378:metabolism; D018053:Arsenites / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D008297:Male; D008930:Mitochondria, Liver / Q000201:enzymology; D025261:Mitochondrial Proton-Translocating ATPases / Q000378:metabolism*; D010084:Oxidation-Reduction; D011117:Polyribonucleotide Nucleotidyltransferase / Q000378:metabolism*; D051381:Rats; D017208:Rats, Wistar
D001149:Arsenates; D018053:Arsenites; D000244:Adenosine Diphosphate; D000255:Adenosine Triphosphate; D011117:Polyribonucleotide Nucleotidyltransferase; D025261:Mitochondrial Proton-Translocating ATPases; D053501:Arsenite Transporting ATPases; D005978:Glutathione