Effect of amino acid substitutions on ArsI activity. (A) MAs(III) demethylation by purified wild type and mutants ArsI was determined. The reaction was initiated by addition of MAs(III), and activity determined by HPLC-ICP-MS, as described in Materials and Methods. (B) MAs(III) demethylation by wild type ArsI and mutants is expressed as a percentage of total arsenic. Demethylation was assayed in mixture of 5 μM MAs(III), 1 μM purified ArsI, 3 mM TCEP, 1 mM cysteine, and 0.1 mM (NH4)2Fe(SO4)2·6H2O in 50 mM MOPS buffer, pH 7.2, at 30 °C. Samples were withdrawn after 60 min, the reaction terminated by addition of 6% (v/v) H2O2, final concentration. Arsenic species were analyzed by HPLC-ICP-MS. The data are the means ± SE (n = 3).