title: Arsenic(III) species inhibit oxidative protein folding in vitro.
abstract: The success of arsenic trioxide in the treatment of acute promyelocytic leukemia has renewed interest in the cellular targets of As(III) species. The effects of arsenicals are usually attributed to their ability to bind vicinal thiols or thiol selenols in prefolded proteins thereby compromising cellular function. The present studies suggest an additional, more pleiotropic, contribution to the biological effects of arsenicals. As(III) species, by avid coordination to the cysteine residues of unfolded reduced proteins, can compromise protein folding pathways. Three representative As(III) compounds (arsenite, monomethylarsenous acid (MMA), and an aryl arsenical (PSAO)) have been tested with three reduced secreted proteins (lysozyme, ribonuclease A, and riboflavin binding protein (RfBP)). Using absorbance, fluorescence, and pre-steady-state methods, we show that arsenicals bind tightly to low micromolar concentrations of these unfolded proteins with stoichiometries of 1 As(III) per 2 thiols for MMA and PSAO and 1 As(III) for every 3 thiols with arsenite. Arsenicals, at 10 microM, strongly disrupt the oxidative folding of RfBP even in the presence of 5 mM reduced glutathione, a competing ligand for As(III) species. MMA catalyzes the formation of amyloid-like monodisperse fibrils using reduced RNase. These in vitro data show that As(III) species can slow, or even derail, protein folding pathways. In vivo, the propensity of As(III) species to bind to unfolded cysteine-containing proteins may contribute to oxidative and protein folding stresses that are prominent features of the cellular response to arsenic exposure.
authors.affiliation: Department of Chemistry and Biochemistry, University of Delaware, Newark, Delaware 19716, USA.
authors.forename: Danny
authors.initials: D
authors.lastname: Ramadan
authors.forename: Pumtiwitt C
authors.initials: PC
authors.lastname: Rancy
authors.forename: Radhika P
authors.initials: RP
authors.lastname: Nagarkar
authors.forename: Joel P
authors.initials: JP
authors.lastname: Schneider
authors.forename: Colin
authors.initials: C
authors.lastname: Thorpe
mesh_terms: D000818:Animals; D001151:Arsenic / Q000378:metabolism* / Q000494:pharmacology*; D018053:Arsenites / Q000378:metabolism / Q000494:pharmacology; D001665:Binding Sites; D002417:Cattle; D002645:Chickens; D004220:Disulfides / Q000737:chemistry; D004229:Dithiothreitol / Q000494:pharmacology; D004926:Escherichia coli / Q000235:genetics; D005978:Glutathione / Q000494:pharmacology; D006801:Humans; D007700:Kinetics; D026901:Membrane Transport Proteins / Q000737:chemistry / Q000378:metabolism; D008956:Models, Chemical; D009113:Muramidase / Q000378:metabolism; D010084:Oxidation-Reduction; D011485:Protein Binding; D019704:Protein Disulfide-Isomerases / Q000378:metabolism; D017510:Protein Folding / Q000187:drug effects*; D011506:Proteins / Q000378:metabolism*; D012259:Ribonuclease, Pancreatic / Q000302:isolation & purification / Q000378:metabolism / Q000648:ultrastructure
chemical_list: D018053:Arsenites; D004220:Disulfides; D026901:Membrane Transport Proteins; D011506:Proteins; C019079:riboflavin-binding protein; D012259:Ribonuclease, Pancreatic; D009113:Muramidase; D019704:Protein Disulfide-Isomerases; D005978:Glutathione; C015001:arsenite; D001151:Arsenic; D004229:Dithiothreitol