Reorientation of the Methyl Group in MAs(III) is the Rate-Limiting Step in the ArsM As(III) S-Adenosylmethionine Methyltransferase Reaction.

The most common biotransformation of trivalent inorganic arsenic (As(III)) is methylation to mono-, di-, and trimethylated species. Methylation is catalyzed by As(III) S-adenosylmethionine (SAM) methyltransferase (termed ArsM in microbes and AS3MT in animals). Methylarsenite (MAs(III)) is both the product of the first methylation step and the substrate of the second methylation step. When the rate of the overall methylation reaction was determined with As(III) as the substrate, the first methylation step was rapid, whereas the second methylation step was slow. In contrast, when MAs(III) was used as the substrate, the rate of methylation was as fast as the first methylation step when As(III) was used as the substrate. These results indicate that there is a slow conformational change between the first and second methylation steps. The structure of CmArsM from the thermophilic alga Cyanidioschyzon merolae sp. 5508 was determined with bound MAs(III) at 2.27 Å resolution. The methyl group is facing the solvent, as would be expected when MAs(III) is bound as the substrate rather than facing the SAM-binding site, as would be expected for MAs(III) as a product. We propose that the rate-limiting step in arsenic methylation is slow reorientation of the methyl group from the SAM-binding site to the solvent, which is linked to the conformation of the side chain of a conserved residue Tyr70

Reorientation of the Methyl Group in MAs(III) is the Rate-Limiting Step in the ArsM As(III) SAdenosylmethionine Methyltransferase Reaction

The most common biotransformation of trivalent inorganic arsenic (As(III)) is methylation to mono-, di-, and trimethylated species. Methylation is catalyzed by As(III) S-adenosylmethionine (SAM) methyltransferase (termed ArsM in microbes and AS3MT in animals). Methylarsenite (MAs(III)) is both the product of the first methylation step and the substrate of the second methylation step. When the rate of the overall methylation reaction was determined with As(III) as the substrate, the first methylation step was rapid, whereas the second methylation step was slow. In contrast, when MAs(III) was used as the substrate, the rate of methylation was as fast as the first methylation step when As(III) was used as the substrate. These results indicate that there is a slow conformational change between the first and second methylation steps. The structure of CmArsM from the thermophilic alga Cyanidioschyzon merolae sp. 5508 was determined with bound MAs(III) at 2.27 Å resolution. The methyl group is facing the solvent, as would be expected when MAs(III) is bound as the substrate rather than facing the SAM-binding site, as would be expected for MAs(III) as a product. We propose that the rate-limiting step in arsenic methylation is slow reorientation of the methyl group from the SAM-binding site to the solvent, which is linked to the conformation of the side chain of a conserved residue Tyr70

Reorientation of the Methyl Group in MAs(III) is the Rate-Limiting Step in the ArsM As(III) S-Adenosylmethionine Methyltransferase Reaction.

The most common biotransformation of trivalent inorganic arsenic (As(III)) is methylation to mono-, di-, and trimethylated species. Methylation is catalyzed by As(III) S-adenosylmethionine (SAM) methyltransferase (termed ArsM in microbes and AS3MT in animals). Methylarsenite (MAs(III)) is both the product of the first methylation step and the substrate of the second methylation step. When the rate of the overall methylation reaction was determined with As(III) as the substrate, the first methylation step was rapid, whereas the second methylation step was slow. In contrast, when MAs(III) was used as the substrate, the rate of methylation was as fast as the first methylation step when As(III) was used as the substrate. These results indicate that there is a slow conformational change between the first and second methylation steps. The structure of CmArsM from the thermophilic alga Cyanidioschyzon merolae sp. 5508 was determined with bound MAs(III) at 2.27 Å resolution. The methyl group is facing the solvent, as would be expected when MAs(III) is bound as the substrate rather than facing the SAM-binding site, as would be expected for MAs(III) as a product. We propose that the rate-limiting step in arsenic methylation is slow reorientation of the methyl group from the SAM-binding site to the solvent, which is linked to the conformation of the side chain of a conserved residue Tyr70

The Structure of an As(III) <i>S</i>‑Adenosylmethionine Methyltransferase with 3‑Coordinately Bound As(III) Depicts the First Step in Catalysis

Arsenic is a ubiquitous environmental toxic substance and a Class 1 human carcinogen. Arsenic methylation by the enzyme As­(III) <i>S</i>-adenosylmethionine (SAM) methyltransferase (ArsM in microbes or AS3MT in animals) detoxifies As­(III) in microbes but transforms it into more toxic and potentially more carcinogenic methylated species in humans. We previously proposed a reaction pathway for ArsM/AS3MT that involves initial 3-coordinate binding of As­(III). To date, reported structures have had only 2-coordinately bound trivalent arsenicals. Here we report a crystal structure of CmArsM from <i>Cyanidioschyzon</i> sp.5508 in which As­(III) is 3-coordinately bound to three conserved cysteine residues with a molecule of the product <i>S</i>-adenosyl-l-homocysteine bound in the SAM binding site. We propose that this structure represents the first step in the catalytic cycle. In a previously reported SAM-bound structure, a disulfide bond is formed between two conserved cysteine residues. Comparison of these two structures indicates that there is a conformational change in the N-terminal domain of CmArsM that moves a loop to allow formation of the 3-coordinate As­(III) binding site. We propose that this conformational change is an initial step in the As­(III) SAM methyltransferase catalytic cycle

Structure of the ArsI C–As Lyase: Insights into the Mechanism of Degradation of Organoarsenical Herbicides and Growth Promoters

Arsenic is a ubiquitous and carcinogenic environmental element that enters the biosphere primarily from geochemical sources, but also through anthropogenic activities. Microorganisms play an important role in the arsenic biogeochemical cycle by biotransformation of inorganic arsenic into organic arsenicals and vice versa. ArsI is a microbial non-heme, ferrous-dependent dioxygenase that transforms toxic methylarsenite [MAs(III)] to less toxic and carcinogenic inorganic arsenite [As(III)] by C-As bond cleavage. An ArsI ortholog, TcArsI, from the thermophilic bacterium Thermomonospora curvata was expressed, purified, and crystallized. The structure was solved in both the apo form and with Ni(II), Co(II), or Fe(III). The MAs(III) binding site is a vicinal cysteine pair in a flexible loop. A structure with the loop occupied with β-mercaptoethanol mimics binding of MAs(III). The structure of a mutant protein (Y100H/V102F) was solved in two different crystal forms with two other orientations of the flexible loop. These results suggest that a loop-gating mechanism controls the catalytic reaction. In the ligand-free open state, the loop is exposed to solvent, where it can bind MAs(III). The loop moves toward the active site, where it forms a closed state that orients the C-As bond for dioxygen addition and cleavage. Elucidation of the enzymatic mechanism of this unprecedented C-As lyase reaction will enhance our understanding of recycling of environmental organoarsenicals

Crystal structure of 4-Amino-3-(4′-chlorophenyl)-4-H-[1,2,4]-triazolo-5-thiol

The crystal structure of 4-amino-3-(4′-chlorophenyl)-4H-[1,2,4]-triazolo-5-thiol (ACPTT) was determined by crystallographic methods. There are two crystallographically independent molecules in the asymmetric unit and are related by pseudo inversion symmetry with each other. A number of C-H…N and N-H…N types of intermolecular interactions stabilize the molecules in the unit cell in addition to van der Waals forces

A disulfide-bond cascade mechanism for arsenic(III) S-adenosylmethionine methyltransferase

Methylation of the toxic metalloid arsenic is widespread in nature. Members of every kingdom have arsenic(III) S-adenosylmethionine (SAM) methyltransferase enzymes, which are termed ArsM in microbes and AS3MT in animals, including humans. Trivalent arsenic(III) is methylated up to three times to form methylarsenite [MAs(III)], dimethylarsenite [DMAs(III)] and the volatile trimethylarsine [TMAs(III)]. In microbes, arsenic methylation is a detoxification process. In humans, MAs(III) and DMAs(III) are more toxic and carcinogenic than either inorganic arsenate or arsenite. Here, new crystal structures are reported of ArsM from the thermophilic eukaryotic alga Cyanidioschyzon sp. 5508 (CmArsM) with the bound aromatic arsenicals phenylarsenite [PhAs(III)] at 1.80 Å resolution and reduced roxarsone [Rox(III)] at 2.25 Å resolution. These organoarsenicals are bound to two of four conserved cysteine residues: Cys174 and Cys224. The electron density extends the structure to include a newly identified conserved cysteine residue, Cys44, which is disulfide-bonded to the fourth conserved cysteine residue, Cys72. A second disulfide bond between Cys72 and Cys174 had been observed previously in a structure with bound SAM. The loop containing Cys44 and Cys72 shifts by nearly 6.5 Å in the arsenic(III)-bound structures compared with the SAM-bound structure, which suggests that this movement leads to formation of the Cys72-Cys174 disulfide bond. A model is proposed for the catalytic mechanism of arsenic(III) SAM methyltransferases in which a disulfide-bond cascade maintains the products in the trivalent state

Reorientation of the Methyl Group in MAs(III) is the Rate-Limiting Step in the ArsM As(III) <i>S</i>‑Adenosylmethionine Methyltransferase Reaction

The most common biotransformation of trivalent inorganic arsenic (As­(III)) is methylation to mono-, di-, and trimethylated species. Methylation is catalyzed by As­(III) <i>S</i>-adenosylmethionine (SAM) methyltransferase (termed ArsM in microbes and AS3MT in animals). Methylarsenite (MAs­(III)) is both the product of the first methylation step and the substrate of the second methylation step. When the rate of the overall methylation reaction was determined with As­(III) as the substrate, the first methylation step was rapid, whereas the second methylation step was slow. In contrast, when MAs­(III) was used as the substrate, the rate of methylation was as fast as the first methylation step when As­(III) was used as the substrate. These results indicate that there is a slow conformational change between the first and second methylation steps. The structure of CmArsM from the thermophilic alga Cyanidioschyzon merolae sp. 5508 was determined with bound MAs­(III) at 2.27 Å resolution. The methyl group is facing the solvent, as would be expected when MAs­(III) is bound as the substrate rather than facing the SAM-binding site, as would be expected for MAs­(III) as a product. We propose that the rate-limiting step in arsenic methylation is slow reorientation of the methyl group from the SAM-binding site to the solvent, which is linked to the conformation of the side chain of a conserved residue Tyr70

Functional and structural characterization of AntR, an Sb(III) responsive transcriptional repressor.

The ant operon of the antimony-mining bacterium Comamonas testosterone JL40 confers resistance to Sb(III). The operon is transcriptionally regulated by the product of the first gene in the operon, antR. AntR is a member of ArsR/SmtB family of metal/metalloid-responsive repressors resistance. We purified and characterized C. testosterone AntR and demonstrated that it responds to metalloids in the order Sb(III)&nbsp;=&nbsp;methylarsenite (MAs(III) &gt;&gt; As(III)). The protein was crystallized, and the structure was solved at 2.1&nbsp;Å resolution. The homodimeric structure of AntR adopts a classical ArsR/SmtB topology architecture. The protein has five cysteine residues, of which Cys103a from one monomer and Cys113b from the other monomer, are proposed to form one Sb(III) binding site, and Cys113a and Cys103b forming a second binding site. This is the first report of the structure and binding properties of a transcriptional repressor with high selectivity for environmental antimony