Adventitious Arsenate Reductase Activity of the Catalytic Domain of the Human Cdc25B and Cdc25C Phosphatases
journal contributionposted on 02.02.2010, 00:00 by Hiranmoy Bhattacharjee, Ju Sheng, A. Abdul Ajees, Rita Mukhopadhyay, Barry P. Rosen
A number of eukaryotic enzymes that function as arsenate reductases are homologues of the catalytic domain of the human Cdc25 phosphatase. For example, the Leishmania major enzyme LmACR2 is both a phosphatase and an arsenate reductase, and its structure bears similarity to the structure of the catalytic domain of human Cdc25 phosphatase. These reductases contain an active site C-X5-R signature motif, where C is the catalytic cysteine, the five X residues form a phosphate binding loop, and R is a highly conserved arginine, which is also present in human Cdc25 phosphatases. We therefore investigated the possibility that the three human Cdc25 isoforms might have adventitious arsenate reductase activity. The sequences for the catalytic domains of Cdc25A, -B, and -C were cloned individually into a prokaryotic expression vector, and their gene products were purified from a bacterial host using nickel affinity chromatography. While each of the three Cdc25 catalytic domains exhibited phosphatase activity, arsenate reductase activity was observed only with Cdc25B and -C. These two enzymes reduced inorganic arsenate but not methylated pentavalent arsenicals. Alteration of either the cysteine and arginine residues of the Cys-X5-Arg motif led to the loss of both reductase and phosphatase activities. Our observations suggest that Cdc25B and -C may adventitiously reduce arsenate to the more toxic arsenite and may also provide a framework for identifying other human protein tyrosine phosphatases containing the active site Cys-X5-Arg loop that might moonlight as arsenate reductases.
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prokaryotic expression vectorenzyme LmACR 2Cdc 25 isoformsCdc 25BCdc 25 phosphataseAdventitious Arsenate Reductase ActivityCdc 25 phosphatasesnickel affinity chromatographymethylated pentavalent arsenicalsX residues form25ACdc 25C PhosphatasesA numberHuman Cdc 25Bphosphate binding loopprotein tyrosine phosphatasesarsenate reductasesarsenate reductase activity