Properties of the arsenate reductase of plasmid R773.
Resistance to toxic oxyanions in Escherichia coli is conferred by the ars operon carried on plasmid R773. The gene products of this operon catalyze extrusion of antimonials and arsenicals from cells of E. coli, thus providing resistance to those toxic oxyanions. In addition, resistance to arsenate is conferred by the product of the arsC gene. In this report, purified ArsC protein was shown to catalyze reduction of arsenate to arsenite. The enzymatic activity of the ArsC protein required glutaredoxin as a source of reducing equivalents. Other reductants, including glutathione and thioredoxin, were not effective electron donors. A spectrophotometric assay was devised in which arsenate reduction was coupled to NADPH oxidation. The results obtained with the coupled assay corresponded to those found by direct reduction of radioactive arsenate to arsenite. The only substrate of the reaction was arsenate (Km = 8 mM); other oxyanions including phosphate, sulfate, and antimonate were not reduced. Phosphate and sulfate were weak inhibitors, while the product, arsenite, was a stronger inhibitor (Ki = 0.1 mM). Arsenate reductase activity exhibited a pH optimum of 6.3-6.8. These results indicate that the ArsC protein is a novel reductase, and elucidation of its enzymatic mechanism should be of interest.
Gladysheva
T B
TB
Department of Biochemistry, Wayne State University School of Medicine, Detroit, Michigan 48201.
Oden
K L
KL
Rosen
B P
BP
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D053501:Arsenite Transporting ATPases; D004926:Escherichia coli / Q000235:genetics; D016623:Ion Pumps; D007700:Kinetics; D009097:Multienzyme Complexes; D010084:Oxidation-Reduction; D010957:Plasmids; D013379:Substrate Specificity
D016623:Ion Pumps; D009097:Multienzyme Complexes; D000251:Adenosine Triphosphatases; D053501:Arsenite Transporting ATPases