Methylation of Arsenic by Recombinant Human Wild-Type Arsenic (+3 Oxidation State) Methyltransferase and its Methionine 287 Threonine (M287T) Polymorph: Role of Glutathione

摘要

Arsenic (+3 oxidation state) methyltransferase (AS3MT) is the key enzyme in the pathway for methylation of arsenicals. A common polymorphism in the AS3MT gene that replaces a threonyl residue in position 287 with a methionyl residue (AS3MT/M287T) occurs at a frequency of about 10% among populations worldwide. Here, we compared catalytic properties of recombinant human wild-type (wt) AS3MT and AS3MT/M287T in reaction mixtures containing S-adenosylmethionine, arsenite (iAs^III) or methylarsonous acid (MAs^III) as substrates and endogenous or synthetic reductants, including glutathione (GSH), a thioredoxin reductase (TR)/thioredoxin (Trx)/NADPH reducing system, or tris (2-carboxyethyl) phosphine hydrochloride (TCEP). With either TR/Trx/NADPH or TCEP, wtAS3MT or AS3MT/M287T catalyzed conversion of iAs^III to MAs^III, methylarsonic acid (MAs^V), dimethylarsinous acid (DMAs^III), and dimethylarsinic acid (DMAs^V); MAs^III was converted to DMAs^III and DMAs^V. Although neither enzyme required GSH to support methylation of iAs^III or MAs^III, addition of 1 mM GSH decreased Km and increased Vmax estimates for either substrate in reaction mixtures containing TR/Trx/NADPH. Without GSH, Vmax and Km values were significantly lower for AS3MT/M287T than for wtAS3MT. In the presence of 1 mM GSH, significantly more DMAs^III was produced from iAs^III in reactions catalyzed by the M287T variant than in wtAS3MT-catalyzed reactions. Thus, 1 mM GSH modulates AS3MT activity, increasing both methylation rates and yield of DMAs^III. AS3MT genotype exemplified by differences in regulation of wtAS3MT and AS3MT/M287T-catalyzed reactions by GSH may contribute to differences in the phenotype for arsenic methylation and, ultimately, to differences in the disease susceptibility in individuals chronically exposed to inorganic arsenic.