Mechanism of thiol-supported arsenate reduction mediated by phosphorolytic-arsenolytic enzymes: I. The role of arsenolysis.
Several mammalian enzymes catalyzing the phosphorolytic-arsenolytic cleavage of their substrates (thus yielding arsenylated metabolites) have been shown to facilitate reduction of arsenate (AsV) to the more toxic arsenite (AsIII) in presence of their substrate and a thiol. These include purine nucleoside phosphorylase (PNP), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), and glycogen phosphorylase-a (GPa). In this work, we tested further enzymes, the bacterial phosphotransacetylases (PTAs) and PNP, for AsV reduction. The PTAs, which arsenolytically cleave acetyl-CoA producing acetyl-arsenate, were compared with GAPDH, which can also form acetyl-arsenate by arsenolysis of its nonphysiological substrate, acetyl-phosphate. As these enzymes also mediated AsV reduction, we can assert that facilitation of thiol-dependent AsV reduction may be a general property of enzymes that catalyze phosphorolytic-arsenolytic reactions. Because with all such enzymes arsenolysis is obligatory for AsV reduction, we analyzed the relationship between these two processes in presence of various thiol compounds, using PNP. Although no thiol influenced the rate of PNP-catalyzed arsenolysis, all enhanced the PNP-mediated AsV reduction, albeit differentially. Furthermore, the relative capacity of thiols to support AsV reduction mediated by PNP, GPa, PTA, and GAPDH apparently depended on the type of arsenylated metabolites (i.e., arsenate ester or anhydride) produced by these enzymes. Importantly, AsV reduction by both acetyl-arsenate-producing enzymes (i.e., PTA and GAPDH) exhibited striking similarities in responsiveness to various thiols, thus highlighting the role of arsenylated metabolite formation. This observation, together with the finding that PNP-mediated AsV reduction lags behind the PNP-catalyzed arsenolysis lead to the hypothesis that arsenolytic enzymes promote reduction of AsV by forming arsenylated metabolites which are more reducible to AsIII by thiols than inorganic AsV. This hypothesis is evaluated in the adjoining paper.
Németi
Balázs
B
Department of Pharmacology and Pharmacotherapy, Toxicology Section, University of Pécs, Medical School, Pécs 7624, Hungary.
Gregus
Zoltán
Z
D000105:Acetyl Coenzyme A / Q000378:metabolism; D000818:Animals; D001149:Arsenates / Q000378:metabolism*; D018053:Arsenites / Q000378:metabolism*; D001426:Bacterial Proteins / Q000378:metabolism*; D002417:Cattle; D004229:Dithiothreitol / Q000378:metabolism; D005978:Glutathione / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000378:metabolism*; D024981:Glycogen Phosphorylase / Q000378:metabolism*; D007288:Inosine / Q000378:metabolism; D007700:Kinetics; D008623:Mercaptoethanol / Q000378:metabolism; D008956:Models, Chemical; D010084:Oxidation-Reduction; D010709:Phosphate Acetyltransferase / Q000378:metabolism*; D011683:Purine-Nucleoside Phosphorylase / Q000378:metabolism*; D011817:Rabbits; D017670:Sodium Compounds / Q000378:metabolism*; D004113:Succimer / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D014494:Unithiol / Q000378:metabolism
D001149:Arsenates; D018053:Arsenites; D001426:Bacterial Proteins; D017670:Sodium Compounds; D013438:Sulfhydryl Compounds; D014494:Unithiol; C017947:sodium arsenite; D007288:Inosine; D008623:Mercaptoethanol; D000105:Acetyl Coenzyme A; C009277:sodium arsenate; D004113:Succimer; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D010709:Phosphate Acetyltransferase; D024981:Glycogen Phosphorylase; D011683:Purine-Nucleoside Phosphorylase; D005978:Glutathione; D004229:Dithiothreitol