New mechanisms of bacterial arsenic resistance.
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ABSTRACT:
SUBMITTER: Yang HC
PROVIDER: S-EPMC6138428 | BioStudies | 2016-01-01
REPOSITORIES: biostudies
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Project description:Arsenic is the most pervasive environmental toxic substance. As a consequence of its ubiquity, nearly every organism has genes for resistance to inorganic arsenic. In bacteria these genes are found largely in bacterial arsenic resistance (ars) operons. Recently a parallel pathway for synthesis and degradation of methylated arsenicals has been identified. The arsM gene product encodes the ArsM (AS3MT in animals) As(iii) S-adenosylmethionine methyltransferase that methylates inorganic trivalent arsenite in three sequential steps to methylarsenite MAs(iii), dimethylarsenite (DMAs(iii) and trimethylarsenite (TMAs(iii)). MAs(iii) is considerably more toxic than As(iii), and we have proposed that MAs(iii) was a primordial antibiotic. Under aerobic conditions these products are oxidized to nontoxic pentavalent arsenicals, so that methylation became a detoxifying pathway after the atmosphere became oxidizing. Other microbes have acquired the ability to regenerate MAs(v) by reduction, transforming it again into toxic MAs(iii). Under this environmental pressure, MAs(iii) resistances evolved, including the arsI, arsH and arsP genes. ArsI is a C-As bond lyase that demethylates MAs(iii) back to less toxic As(iii). ArsH re-oxidizes MAs(iii) to MAs(v). ArsP actively extrudes MAs(iii) from cells. These proteins confer resistance to this primitive antibiotic. This oscillation between MAs(iii) synthesis and detoxification is an essential component of the arsenic biogeocycle.
2016-01-01 | S-EPMC5117644 | BioStudies
Project description:Microbes play a critical role in the global arsenic biogeocycle. Most studies have focused on redox cycling of inorganic arsenic in bacteria and archaea. The parallel cycles of organoarsenical biotransformations are less well characterized. Here we describe organoarsenical biotransformations in the environmental microbe Shewanella putrefaciens. Under aerobic growth conditions, S. putrefaciens reduced the herbicide MSMA (methylarsenate or MAs(V)) to methylarsenite (MAs(III)). Even though it does not contain an arsI gene, which encodes the ArsI C-As lyase, S. putrefaciens demethylated MAs(III) to As(III). It cleaved the C-As bond in aromatic arsenicals such as the trivalent forms of the antimicrobial agents roxarsone (Rox(III)), nitarsone (Nit(III)) and phenylarsenite (PhAs(III)), which have been used as growth promoters for poultry and swine. S. putrefaciens thiolated methylated arsenicals, converting MAs(V) into the more toxic metabolite monomethyl monothioarsenate (MMMTAs(V)), and transformed dimethylarsenate (DMAs(V)) into dimethylmonothioarsenate (DMMTAs(V)). It also reduced the nitro groups of Nit(V), forming p-aminophenyl arsenate (p-arsanilic acid or p-AsA(V)), and Rox(III), forming 3-amino-4-hydroxybenzylarsonate (3A4HBzAs(V)). Elucidation of organoarsenical biotransformations by S. putrefaciens provides a holistic appreciation of how these environmental pollutants are degraded.
2016-01-01 | S-EPMC4984541 | BioStudies
Project description:Environmental organoarsenicals are produced by microorganisms and are introduced anthropogenically as herbicides and antimicrobial growth promoters for poultry and swine. Nearly every prokaryote has an ars (arsenic resistance) operon, and some have an arsH gene encoding an atypical flavodoxin. The role of ArsH in arsenic resistance has been unclear. Here we demonstrate that ArsH is an organoarsenical oxidase that detoxifies trivalent methylated and aromatic arsenicals by oxidation to pentavalent species. Escherichia coli, which does not have an arsH gene, is very sensitive to the trivalent forms of the herbicide monosodium methylarsenate [MSMA or MAs(V)] and antimicrobial growth promoter roxarsone [Rox(V)], as well as to phenylarsenite [PhAs(III), also called phenylarsine oxide or PAO]. Pseudomonas putida has two chromosomally encoded arsH genes and is highly resistant to the trivalent forms of these organoarsenicals. A derivative of P. putida with both arsH genes deleted is sensitive to MAs(III), PhAs(III) or Rox(III). P.?putida?arsH expressed in E.?coli conferred resistance to each trivalent organoarsenical. Cells expressing PpArsH oxidized the trivalent organoarsenicals. PpArsH was purified, and the enzyme in vitro similarly oxidized the trivalent organoarsenicals. These results suggest that ArsH catalyzes a novel biotransformation that confers resistance to environmental methylated and aromatic arsenicals.
2015-01-01 | S-EPMC4447588 | BioStudies
Project description:Arsenic is the most widespread environmental toxin. Substantial amounts of pentavalent organoarsenicals have been used as herbicides, such as monosodium methylarsonic acid (MSMA), and as growth enhancers for animal husbandry, such as roxarsone (4-hydroxy-3-nitrophenylarsonic acid) [Rox(V)]. These undergo environmental degradation to more toxic inorganic arsenite [As(III)]. We previously demonstrated a two-step pathway of degradation of MSMA to As(III) by microbial communities involving sequential reduction to methylarsonous acid [MAs(III)] by one bacterial species and demethylation from MAs(III) to As(III) by another. In this study, the gene responsible for MAs(III) demethylation was identified from an environmental MAs(III)-demethylating isolate, Bacillus sp. MD1. This gene, termed arsenic inducible gene (arsI), is in an arsenic resistance (ars) operon and encodes a nonheme iron-dependent dioxygenase with C ? As lyase activity. Heterologous expression of ArsI conferred MAs(III)-demethylating activity and MAs(III) resistance to an arsenic-hypersensitive strain of Escherichia coli, demonstrating that MAs(III) demethylation is a detoxification process. Purified ArsI catalyzes Fe(2+)-dependent MAs(III) demethylation. In addition, ArsI cleaves the C ? As bond in trivalent roxarsone and other aromatic arsenicals. ArsI homologs are widely distributed in prokaryotes, and we propose that ArsI-catalyzed organoarsenical degradation has a significant impact on the arsenic biogeocycle. To our knowledge, this is the first report of a molecular mechanism for organoarsenic degradation by a C ? As lyase.
2014-01-01 | S-EPMC4040596 | BioStudies
Project description:Arsenite [As(III)] and methylarsenite [MAs(III)] are the most toxic inorganic and methylated arsenicals, respectively. As(III) and MAs(III) can be interconverted in the unicellular cyanobacterium Nostoc sp. PCC 7120 (Nostoc), which has both the arsM gene (NsarsM), which is responsible for arsenic methylation, and the arsI gene (NsarsI), which is responsible for MAs(III) demethylation. It is not clear how the cells prevent a futile cycle of methylation and demethylation. To investigate the relationship between arsenic methylation and demethylation, we constructed strains of Escherichia coli AW3110 (?arsRBC) expressing NsarsM or/and NsarsI. Expression of NsarsI conferred MAs(III) resistance through MAs(III) demethylation. Compared to NsArsI, NsArsM conferred higher resistance to As(III) and lower resistance to MAs(III) by methylating both As(III) and MAs(III). The major species found in solution was dimethylarsenate [DMAs(V)]. Co-expression of NsarsM and NsarsI conferred As(III) resistance at levels similar to that with NsarsM alone, although the main species found in solution after As(III) biotransformation was methylarsenate [MAs(V)] rather than DMAs(V). Co-expression of NsarsM and NsarsI conferred a higher level of resistance to MAs(III) than found with expression of NsarsM alone but lower than expression of only NsarsI. Cells co-expressing both genes converted MAs(III) to a mixture of As(III) and DMAs(V). In Nostoc NsarsM is constitutively expressed, while NsarsI is inducible by either As(III) or MAs(III). Thus, our results suggest that at low concentrations of arsenic, NsArsM activity predominates, while NsArsI activity predominates at high concentrations. We propose that coexistence of arsM and arsI genes in Nostoc could be advantageous for several reasons. First, it confers a broader spectrum of resistance to both As(III) and MAs(III). Second, at low concentrations of arsenic, the MAs(III) produced by NsArsM will possibly have antibiotic-like properties and give the organism a competitive advantage. Finally, these results shed light on the role of cyanobacteria in the arsenic biogeochemical cycle.
2017-01-01 | S-EPMC5258700 | BioStudies
Project description: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.
2015-01-01 | S-EPMC4356363 | BioStudies
Project description:Arsenic is the most ubiquitous environmental toxin. Here, we demonstrate that bacteria have evolved the ability to use arsenic to gain a competitive advantage over other bacteria at least twice. Microbes generate toxic methylarsenite (MAs(III)) by methylation of arsenite (As(III)) or reduction of methylarsenate (MAs(V)). MAs(III) is oxidized aerobically to MAs(V), making methylation a detoxification process. MAs(V) is continually re-reduced to MAs(III) by other community members, giving them a competitive advantage over sensitive bacteria. Because generation of a sustained pool of MAs(III) requires microbial communities, these complex interactions are an emergent property. We show that reduction of MAs(V) by Burkholderia sp. MR1 produces toxic MAs(III) that inhibits growth of Escherichia coli in mixed culture. There are three microbial mechanisms for resistance to MAs(III). ArsH oxidizes MAs(III) to MAs(V). ArsI degrades MAs(III) to As(III). ArsP confers resistance by efflux. Cells of E. coli expressing arsI, arsH or arsP grow in mixed culture with Burkholderia sp. MR1 in the presence of MAs(V). Thus MAs(III) has antibiotic properties: a toxic organic compound produced by one microbe to kill off competitors. Our results demonstrate that life has adapted to use environmental arsenic as a weapon in the continuing battle for dominance.
2019-01-01 | S-EPMC6370046 | BioStudies
Project description:Organoarsenicals such as the methylarsenical methylarsenate (MAs(V)) and aromatic arsenicals including roxarsone (4-hydroxy-3-nitrobenzenearsenate or Rox(V)) have been extensively used as an herbicide and growth enhancers in animal husbandry, respectively. They undergo environmental degradation to more toxic inorganic arsenite (As(III)) that contaminates crops and drinking water. We previously identified a bacterial gene (arsI) responsible for aerobic demethylation of methylarsenite (MAs(III)). The gene product, ArsI, is an Fe(II)-dependent extradiol dioxygenase that cleaves the carbon-arsenic (C-As) bond in MAs(III) and in trivalent aromatic arsenicals. The objective of this study was to elucidate the ArsI mechanism. Using isothermal titration calorimetry, we determined the dissociation constants and ligand-to-protein stoichiometry of ArsI for Fe(II), MAs(III), and aromatic phenylarsenite. Using a combination of methods including chemical modification, site-directed mutagenesis, and fluorescent spectroscopy, we demonstrated that amino acid residues predicted to participate in Fe(II)-binding (His5-His62-Glu115) and substrate binding (Cys96-Cys97) are involved in catalysis. Finally, the products of Rox(III) degradation were identified as As(III) and 2-nitrohydroquinone, demonstrating that ArsI is a dioxygenase that incorporates one oxygen atom from dioxygen into the carbon and the other to the arsenic to catalyze cleavage of the C-As bond. These results augment our understanding of the mechanism of this novel C-As lyase.
2017-01-01 | S-EPMC5870903 | BioStudies
Project description:The toxic metalloid arsenic is widely distributed in food, water, and soil. While inorganic arsenic enters the environment primarily from geochemical sources, methylarsenicals either result from microbial biotransformation of inorganic arsenic or are introduced anthropogenically. Methylarsenicals such as monosodium methylarsonic acid (MSMA) have been extensively utilized as herbicides, and aromatic arsenicals such as roxarsone (Rox) are used as growth promoters for poultry and swine. Organoarsenicals are degraded to inorganic arsenic. The toxicological effects of arsenicals depend on their oxidation state, chemical composition, and bioavailability. Here we report that the active forms are the trivalent arsenic-containing species. We constructed a whole-cell biosensor utilizing a modified ArsR repressor that is highly selective toward trivalent methyl and aromatic arsenicals, with essentially no response to inorganic arsenic. The biosensor was adapted for in vitro detection of organoarsenicals using fluorescence anisotropy of ArsR-DNA interactions. It detects bacterial biomethylation of inorganic arsenite both in vivo and in vitro with detection limits of 10(-7) M and linearity to 10(-6) M for phenylarsenite and 5 × 10(-6) M for methylarsenite. The biosensor detects reduced forms of MSMA and roxarsone and offers a practical, low cost method for detecting activate forms and breakdown products of organoarsenical herbicides and growth promoters.
2014-01-01 | S-EPMC3939449 | BioStudies
Project description:Microbial expression of genes for resistance to heavy metals and metalloids is usually transcriptionally regulated by the toxic ions themselves. Arsenic is a ubiquitous, naturally occurring toxic metalloid widely distributed in soil and groundwater. Microbes biotransform both arsenate (As(V)) and arsenite (As(III)) into more toxic methylated metabolites methylarsenite (MAs(III)) and dimethylarsenite (DMAs(III)). Environmental arsenic is sensed by members of the ArsR/SmtB family. The arsR gene is autoregulated and is typically part of an operon that contains other ars genes involved in arsenic detoxification. To date every identified ArsR is regulated by inorganic As(III). Here we described a novel ArsR from Shewanella putrefaciens selective for MAs(III). SpArsR orthologs control expression of two MAs(III) resistance genes, arsP that encodes the ArsP MAs(III) efflux permease, and arsH encoding the ArsH MAs(III) oxidase. SpArsR has two conserved cysteine residues, Cys101 and Cys102. Mutation of either resulted in loss of MAs(III) binding, indicating that they form an MAs(III) binding site. SpArsR can be converted into an As(III)-responsive repressor by introduction of an additional cysteine that allows for three-coordinate As(III) binding. Our results indicate that SpArsR evolved selectivity for MAs(III) over As(III) in order to control expression of genes for MAs(III) detoxification.
2017-01-01 | S-EPMC5653410 | BioStudies