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Arsenic metabolism by microbes in nature and the impacton arsenic remediationShen-Long Tsai1,4, Shailendra Singh1,2,3,4 and Wilfred Chen1
In nature, both prokaryotes and eukaryotes have evolved a
wide spectrum of pathways such as oxidation/reduction,
compartmentalization, exclusion, and immobilization [16] as
the main natural defense mechanisms to arsenic. This review
highlights our current understanding of the biochemistry and
molecular biology involved in these natural arsenic
metabolisms, and some successful examples of engineered
microbes by harnessing these natural mechanisms for effective
remediation.
Addresses1 Department of Chemical and Environmental Engineering,
University of California, Riverside, CA 92521, United States2 Cell Molecular and Developmental Biology Program,
University of California, Riverside, CA 92521, United States3 Current address: One MedImmune Way, Gaithersburg, MD 20878,
United States.4 These authors contributed equally to this work.
Corresponding author: Chen, Wilfred ([email protected])
Current Opinion in Biotechnology 2009, 20:659–667
This review comes from a themed issue on
Chemical biotechnology
Edited by Kazuya Watanabe and George Bennett
Available online 31st October 2009
0958-1669/$ – see front matter
# 2009 Elsevier Ltd. All rights reserved.
DOI 10.1016/j.copbio.2009.09.013
IntroductionArsenic (As) is a natural and ubiquitous element that
presents in many environmental compartments and is
released through various natural processes or by anthro-
pogenic inputs. It is recognized as carcinogenic [1] and
chronic exposure to arsenic results in a wide range of
adverse health effects [2,3]. Depending on the physical–chemical conditions of the environment, some arsenic
compounds can be easily solubilized in water [4] and
taken up by microorganisms, resulting in high levels of
bioavailability [5]. The most notable case was observed in
India and Bangladesh where over 50 million people were
exposed to highly contaminated water or food [6]. There
have been reports of up to 2 mg/kg of arsenic accumulated
in grains [7] and up to 92 mg/kg of arsenic in straws [8].
Arsenic occurs in several oxidation states including
arsenate As(V), arsenite As(III), elemental As(0) and
arsenide As(�III). In natural waters, arsenic is mostly
found in its inorganic forms as trivalent arsenite [As(III)]
or pentavalent arsenate [As(V)] [9]. Among them, As(III)
is generally considered to be more mobile and more toxic
than As(V) [10]. Substitutions for phosphate and sub-
sequent inhibition of oxidative phosphorylation is the
major toxicity of pentavalent As(V) [11]. On the other
hand, the affinity of trivalent As(III) for protein thiols or
vicinal sulfhydryl groups makes them highly toxic. As(III)
also acts as an endocrine disruptor by binding to hormone
receptors and interferes with normal cell signaling [12].
Arsenite-stimulated generation of Reactive Oxygen
Species is known to damage proteins, lipids, and DNA
and is probably the direct cause of the carcinogenicity of
arsenite [10].
Owing to its extreme toxicity, arsenic is ranked number
one on the Environmental Protection Agency’s (EPA)
priority list of drinking water contaminants and effective
from 2006 the maximum contaminant level for arsenic in
drinking water was reduced by the US Environmental
Protection Agency from 50 ppb to 10 ppb. According to
the Natural Resources Defense Council, over 56 million
Americans in the 25 reporting states consume water
containing arsenic at levels presenting a potential fatal
cancer risk. Several treatment technologies have been
applied in laboratory-scale and/or field-scale testing for
the removal of arsenic from waters, such as coagulation,
filtration, ion exchange, adsorption, and reverse osmosis
[13–15]. However, these technologies are either too
expensive or ineffective for low arsenic concentration
treatment. To comply with the current regulatory limit
of 10 ppb would require extensive technological devel-
opments that are highly selective and economically com-
petitive.
In nature, microbes respond to arsenic in a variety of
different ways. Depending on the species of different
microorganisms, the responses could be chelation, com-
partmentalization, exclusion, and immobilization [16].
Understanding the molecular and genetic level of
arsenic metabolism will be, therefore, an important
knowledge base for developing efficient and selective
arsenic bioremediation approaches, which has so far
been considered as a cost-effective and environmental
friendly way for heavy-metal removal. In this review, we
will highlight the natural arsenic metabolism in differ-
ent microbes and their impact on environmental arsenic
contamination. In addition, the potential utility of these
natural metabolisms for arsenic remediation will be
discussed.
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Arsenic in the environmentThe major source of As contamination is from naturally
existing minerals; however, anthropogenic activities have
also contributed extensively [17] (Figure 1). As exists in
several oxidation states (�3, 0, +3, and +5), enabling it to
mobilize under various environmental conditions and
hinders many remediation technologies from efficiently
removing it from water. Under oxidizing conditions,
As(V) is the dominant form at lower pH while As(III)
becomes dominant at higher pH (Figure 1). However, the
uncharged form of As(III) [As(OH)3] becomes dominant
under reducing environments, which is more toxic and
difficult to remove [18]. Nitrate can greatly influence As
cycling by oxidizing ferrous iron to produce As-sorbing
particles [19]. Elemental arsenic is not common and
organic arsines are only found in extremely reducing
environments [20]. A number of microorganisms have
been shown to methylate arsenic giving rise to mono-
methyl, dimethyl, and/or tri-methyl derivatives [21].
These methylated arsines are volatile and are rapidly
released to the atmosphere.
The widespread presence of arsenic has forced different
microorganisms to develop arsenic detoxification machi-
neries. Microorganisms have developed various strategies
to counter-act arsenic toxicity: firstly, active extrusion of
arsenic; secondly, intracellular chelation (in eukaryotes)
by various metal-binding peptides including glutathione
(GSH), phytochelatins (PCs), and metallothioneins
(MTs); thirdly, arsenic transformation to various organic
forms which could be potentially less toxic (Figure 2). In
the forthcoming sections we will discuss in detail about
these mechanisms in prokaryotes and eukaryotes.
Arsenic metabolism by prokaryotesArsenic uptake pathways
Arsenic could potentially act as an electron donor or
acceptor and be part of the electron transport chain in
some bacteria. However, specific uptake transporters
have not evolved because of the extreme toxicity [22].
As(III) and As(V) are typically taken up using the glycerol
and phosphate transporter, respectively, because of their
structure chemical similarities to As(III) and As(V). In E.coli, for example, two phosphate transporters (Pit and Pst)
are used for As(V) uptake, with Pst being the dominant
uptake pathway [23�]. The uncharged As(III) is taken up
by the glycerol transporter GlpF [24], a member of
glycerol channels of the major intrinsic protein (MIP)
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Figure 1
Geo cycling of arsenic.
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family. Mutation in GlpF resulted in As(III)-tolerant
E. coli strains [24]. GlpF homologs have been identified
in Leishmania major [25] or Pseudomonas putida, and are
likely to facilitate As(III) transport across the cell mem-
brane in these species.
Basic detoxification mechanisms
Many Gram-negative and Gram-positive bacteria employ
a similar arsenic resistance mechanism based on the ars
operon (typically arsRDABC) encoded either on the
chromosome or on plasmids [26]. In both cases, there
are two necessary components: a reductase enzyme
(ArsC) for the reduction of As(V) to As(III), which is
subsequently extruded using an As(III) expulsion pump
(ArsB). Additional ars genes have recently been found
suggesting parallel evolution and complex regulations
[27]. The source of reducing power varies among prokar-
yotes; while E. coli employs GSH and glutaredoxin [28],
Arsenic metabolism by microbes in nature Tsai, Singh and Chen 661
Figure 2
Schematic representations of (a) prokaryotes’ and (b) eukaryotes’ processes involved in arsenic metabolism in the environment. In both cases, arsenic
enters the cells through transporters. Arsenate is reduced to arsenite by a reductase, which further extrudes out of the cell by a specific membrane
pump. In eukaryotes, arsenite can also be detoxified by complexation with Cys-rich peptides such as phytochelatins and storage in the vacuole. In
addition, arsenite can serve as an electron donor by oxidation to arsenate. Arsenate can be used as the ultimate electron acceptor during respiration
and inorganic arsenic can also be transformed into organic species in a methylation cascade.
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Staphylococcus aureus utilizes thioredoxin [29�]. During the
reduction step, arsenate binds to a recognition domain
comprising of Arg Residues, resulting in a disulfide bond
between the cysteine residues on ArsC and the reducing
equivalents. Reduction of the disulfide bond via electron
transfer results in As(V) reduction into As(III) [30].
ArsR and ArsD are regulatory components primarily act-
ing as a transcription repressor and regulating the upper
limit for operon activity, respectively [31]. These regu-
latory proteins have extremely high affinity for As(III) and
bind via their cysteine residues, resulting in altered DNA
binding for transcriptional activation [32]. ArsA is an
ATPase that assists ArsB in As(III) efflux by providing
the necessary energy via ATP hydrolysis [33]. Interest-
ingly, the relatively less toxic As(V) is converted to the
more toxic As(III) before efflux; it is possible that the
As(III) efflux system was first evolved under reducing
environments, which was subsequently coupled with
As(V) reduction to accommodate As(V) toxicity once
the earth atmosphere became more oxidized [31].
Arsenite oxidation/reduction
Oxidization of As(III) can be important for arsenic
removal since As(V) is less soluble and is much more
effectively removed by physico-chemical methods [34].
In nature, microorganisms carry out As(III) oxidation
using the enzyme As(III) oxidase, which is classified as
a member of the DMSO reductase family and was only
recently identified and sequenced [35]. Most arsenite
oxidases, like the one (AoxAB) isolated from Hydrogeno-phaga sp. strain NT-14, work as a heterodimer (from the
gene aoxAB) and contain Fe and molybdenum as part of
the catalytic unit [35]. Phylogenetic lineages suggested
that the enzyme had an early origin primarily as a resist-
ance mechanism converting the more toxic As(III) to the
less toxic As(V). However, some chemolithotropic bac-
teria do extract energy from oxidizing arsenite [36].
In addition to the intracellular reduction of As(V) using
the arsenate reductase, arsenate reduction can also be part
of the anaerobic arsenate respiration in some bacteria (e.g.
Shewanella sp. strain ANA-3) [37], where arsenate acts as a
terminal electron acceptor. This respiratory arsenate
reductase (ArrA and ArrB) is membrane-bound like other
members of the electron transport chain [38] and contains
a molybdopterin center in ArrA and a Fe–S center in ArrB.
A Shewanella sp. strain ANA-3 containing a mutation in
the arrAB gene cluster is unable to grow on As(V) [39].
Methylation/demethylation
Methylation is originally thought as a detoxification step;
however, recent literature suggests that not all methyl-
ated arsenic products are less toxic [20]. The primary
mode of arsines and methyl arsenicals generation is As(V)
reduction and subsequent oxidative addition of methyl
groups [40] from various sources such as methyl cobala-
mine in many bacterial systems [41]. Methylated forms of
arsenic are volatile and readily released into the environ-
ment where oxidation might convert them back to the
oxidized form As(V). Very little is known about the
demethylation pathways; however, demethylation of mo-
no-methyl and dimethyl arsenic compounds have been
demonstrated and even the use of methylated arsenicals
as a carbon source is possible [42]. The understanding of
these mechanisms will not only shed light on the arsenic
mobilization but may also open up new horizons in
metabolic pathway engineering to exploit those pathways
for arsenic remediation.
Arsenic efflux machinery
As(III) can either be extruded via an arsenite carrier
protein or via an arsenite efflux pump ArsB. The first
approach exploits the membrane potential for energy
while the latter utilizes the energy provided by the
ATPase ArsA via ATP hydrolysis [43]. The majority of
prokaryote systems employ the ArsA/B system while
some bacteria can suffice only with ArsB. Reduced affi-
nity for As(III) after cysteine residue mutations suggests
that ArsA activation by As(III) occurs via metal-thiolate
complex formed among three cysteine residues and
As(III) [44].
Arsenic metabolism by eukaryotesThe arsenic metabolism by plant cells has recently been
reviewed elaborately elsewhere [45,46��,47]. In this part,
we will only highlight the metabolism of yeast, fungi, and
algae when exposed to arsenic compounds. These would
include the mechanism of arsenic uptake, metabolism,
and efflux.
Arsenic uptake
Arsenic uptake by Saccharomyces cerevisiae occurs through
three different transport systems. The pentavalent
arsenate, because of the similarity to phosphate [48], is
taken up through a phosphate transporter, Pho87p [49]. In
addition, two transporter systems for the trivalent arsenite
have been identified. Similar to bacterial systems,
arsenite is taken up by an aquaglyceroporin Fps1p, a
glycerol transporter [50,51]. Disruption of the FPS1 gene
resulted in a reduction in arsenite uptake, which confirms
the important role of the Fpslp channel for arsenite
uptake [52,53]. However, the FPS1 deletion strain was
still sensitive to arsenite in the absence of glucose
suggesting the existence of an additional transport mech-
anism related to glucose uptake [54]. In 2004, Liu et al.found that a class of hexose permeases (Hxt1p to Hxt1
plus Gal2p) of S. cerevisiae adventitiously catalyzed the
uptake of arsenite [55��]. Arsenite uptake was reduced by
80% in the presence of glucose even when FPS1 was
deleted, confirming that the hexose transporters are
mainly responsible for arsenite uptake. Recently, the
same group demonstrated that a mammalian glucose
662 Chemical biotechnology
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permease GLUT also catalyzed the uptake of arsenite
when heterogeneously expressed in yeast [56].
Arsenic metabolism
Once arsenic enters the cells, a series of detoxification
steps are used to reduce the acute cytotoxic effects. The
most comprehensive mechanism of arsenic tolerance in
yeast is provided by three contiguous gene clusters:
ARR1, ARR2, and ARR3. ARR1 encodes a transcription
factor that regulates the transcription of arsenate
reductase Arr2p and the arsenite extrusion transporter
Arr3p [57�]. After arsenate is transported inside the yeast
cells, arsenate is reduced to arsenite by an arsenate
reductase Arr2p [58]. However, unlike the bacterial
arsenate reductase ArsC (a 141-residue monomer), Arr2p
is a homodimer of two 130-residue monomers. It has been
shown that the yeast gene ARR2 can complement an E.coli strain with a deletion of the chromosomal arsC gene
[58]. In addition, the disruption of ARR2 in S. cerevisiaeeliminated arsenate resistance [59]. Therefore, the resist-
ance of cultured cells for arsenic toxicity has long been
thought to reduce the accumulation of arsenite since no
arsenate efflux transporter has been found so far. To date
Arr2p is still the sole arsenate reductase in eukaryote and
no ARR2 gene has yet been found with the fission yeast S.pombe or other fungi.
Intracellular sequestration
Questions have been raised as to why cells were
designed to reduce arsenate to the more reactive
arsenite, which is at least 100 times more toxic [60].
The answer is that by taking advantage of the chemical
reactivity, arsenite can bind to many intracellular chelat-
ing proteins or peptides containing thiol ligands, such as
GSH, PCs, and MTs to form inactive complexes [61–63].
GSH is a major reservoir of nonprotein thiols [64],
and the availability of GSH is important in arsenate
reduction as well as in arsenite transport into the
vacuoles [65]. Guo et al. showed that overexpression of
the S. cerevisiae GSH1 gene encoding a g-glutamylcys-
teine synthetase (g-ECS), the first enzyme in the GSH
biosynthesis pathway [66], elevated the tolerance and
accumulation of arsenic in Arabidopsis thaliana [67]. MTs
belong to a family of cysteine-rich proteins with the
unique ability to form stable metal-thiolate clusters with
their two metal-binding, cysteine-rich domains [68], and
are the major metal-binding ligands in animals. Although
As-binding MTs have been described in the alga Fucusvesiculosus [69], none have been isolated in bacteria. On
the other hand, PCs are small enzymatically synthesized
cysteine-rich peptides widely found in plants and yeasts,
and have been shown to bind arsenite efficiently
[70,71,72�]. Overexpression of a tobacco PC synthase
in yeast S. cerevisiae resulted in increased tolerance for Cd
and As [73] without any enhancement in accumulation.
However, our lab reported enhanced accumulation of
arsenite by engineered S. cerevisiae expressing the Ara-bidopsis thaliana PC synthase [74].
For some yeasts such as Candida glabrata, extracellular
sulfate is metabolized to sulfide [75], which acts as an
electron donor for arsenate reduction [76]. In some eukar-
yotes, incorporation of sulfide to form a more stable, high-
molecular-weight PC–metal–sulfide complex in the
vacuole has been demonstrated [77–79]. In addition,
the formation of metal sulfide particles in Schizosacchar-omyces pombe and Candida glabrata is also part of their
intracellular detoxification [80,81].
Arsenic resistant via intracellular and extracellular
transport
S. cerevisiae has two different mechanisms to reduce
arsenite cytotoxicity. One is through the arsenite extru-
sion pump Arr3p, which transports the As(III)–GSH
complexes out of the membrane. Overexpression of Arr3p
in yeast results in As(III) tolerance [82], while deletion of
ARR3 results in sensitivity to both As(V) and As(III) [50–52]. In addition to the membrane efflux pump, a second
mechanism of arsenic resistance is via the transport of
GSH-conjugated arsenite into the vacuole [52]. The
Ycf1p protein associated with the vacuolar membrane
is a member of the ABC transporter superfamily that is
responsible for the ATP-dependent transport of a wide
range of GSH-conjugated substrates (such as As(GS)3)
into the vacuole. Both of these mechanisms are essential
for survival at high arsenic concentrations as deletion of
the YCF1 gene results in arsenic hypersensitivity. Further
genetic analyses support the notion that these two path-
ways function in a synergistic fashion as the hypersensi-
tivity of yeast cells to arsenic is additive in a mutant
lacking both genes. While S. cerevisiae transports the
GSH–As complex into the vacuole, S. pombe transports
high-molecular PC–Cd–S complexes into the vacuole via
the Hmt1 transporter [83].
Engineered microbes for arsenic remediationThe use of engineered microbes as selective biosorbents
is an attractive green technology for the low-cost and
efficient removal of arsenic [74]. Although efforts have
been reported in engineering microbes for the removal of
cadmium or mercury by expressing metal-binding pep-
tides such as human MTs [84,85] or synthetic peptides
[86,87], the relatively low specificity and affinity of these
peptides for arsenic make them ineffective for arsenic
remediation. Development of an arsenic accumulating
microbe should comprise the ability to firstly, modify the
naturally existing defense mechanisms and secondly,
develop novel or hybrid pathways into one easily manipu-
lated microorganism.
One of the earliest examples of engineering arsenic
accumulation was demonstrated in plants. The bacterial
enzymes ArsC (arsenate reductase) and g-ECS (GSH
Arsenic metabolism by microbes in nature Tsai, Singh and Chen 663
www.sciencedirect.com Current Opinion in Biotechnology 2009, 20:659–667
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synthase) were expressed in A. thaliana, resulting in the
accumulation of As(V) as GSH–As complexes [88��]. A
similar effort was subsequently reported by expressing
the yeast YCF1 in A. thaliana for enhanced As storage in
the vacuole [89]. These reports open up the possibility of
engineering metabolisms and pathways for arsenic
sequestration. On the basis of these early examples,
similar efforts have been demonstrated with engineered
microbes. In one case, the PC synthase from A. thalianawas expressed in E. coli [90�]. This engineered strain
produced PC when exposed to different forms of arsenic,
leading to moderate levels of arsenic accumulation. How-
ever, the level of GSH, a key PC precursor, became
limiting for higher level of PC production and arsenic
accumulation. Our lab has recently expressed the PC
synthase from S. pombe (SpPCS) in E. coli, resulting in
higher As accumulation [98]. PC production was further
increased by coexpressing a feedback desensitized g-
glutamylcysteine synthetase (GshI*), resulting in higher
PC levels and As accumulation. The significantly
increased PC levels were exploited further by coexpres-
sing an arsenic transporter GlpF, leading to an additional
1.5-fold higher As accumulation. These engineering steps
were finally combined in an arsenic efflux deletion E. colistrain to achieve the highest reported arsenic accumu-
lation in E. coli of 16.8 mmol/g cells.
Naturally, sulfur reducing bacteria are used for As(V)
precipitation by the formation of insoluble sulfide com-
plex with H2S [91]. Metabolic engineering approaches
have been utilized for intracellular production of H2S in
bacteria, leading to higher cadmium accumulation [92].
Our lab has recently engineered a yeast strain coexpres-
sing AtPCS and cysteine desulfhydrase, an aminotrans-
ferase that converts cysteine into hydrogen sulfide under
aerobic condition, to elevate the accumulation of arsenic
by the formation of PC–metal–sulfide complexes (Tsai,
2009, unpublished).
The use of resting cells as a high-affinity biosorbent for
arsenic removal has also been exploited. By expressing
AtPCS in S. cerevisiae, which naturally has a higher level of
GSH, the engineered yeast strain accumulated high
levels of arsenic and was effective in removing arsenic
in resting cell cultures [74]. However, the utility of PC-
producing cells for biosorption necessitates the use of
zinc for PC induction, making it difficult to implement
in practice. On the other hand, specific arsenic accumu-
lation was achieved in E. coli cells by overexpressing the
arsenic-specific regulatory protein ArsR. Resting cells
expressing ArsR were effective in removing 50 ppb of
As(III) within one hour [93]. The concept of resting cell
sorbents has been extended to the use of a naturally
occurring As-binding MT [94��]. Singh and coworkers
developed an engineered E. coli strain expressing the
fMT from F. vesiculosus [69] isolated from an arsenic-
contaminated site. When the arsenite-specific transporter
GlpF was co-overexpressed with fMT, the engineered
E. coli accumulated arsenic at high levels even in the
presence of 10-fold excess amounts of competing heavy
metals [94��]. Resting cells were able to completely
remove 35 ppb of As(III) within 20 min, making this an
attractive low-cost option for arsenic remediation.
New irrational approaches such as directed evolution,
genome shuffling, and metagenomic studies can be used
for developing new arsenic resistant pathways that are
suitable for arsenic remediation [95]. This was demon-
strated by the modification of an arsenic resistance operon
using DNA shuffling [96��]. Cells expressing the opti-
mized operon grew in 0.5 M arsenate, a 40-fold increase in
resistance. Along the same line, Chauhan and coworkers
constructed a metagenomic library from an industrial
effluent treatment plant sludge, and identified a novel
As(V) resistance gene (arsN) encoding a protein similar to
acetyltransferases. Overexpression of ArsN led to higher
arsenic resistance in E. coli [97]. These examples high-
light the possibility to combine both natural and unna-
tural pathways for hyperarsenic accumulation.
ConclusionArsenic contamination is a major global problem and local
geochemical cycles have been intensified by irresponsi-
ble industrial and mining activities. Fortunately, many
microorganisms have already evolved mechanisms to
cope with this environmental challenge. The fundamen-
tal understanding of the biochemistry and metabolic
pathways involved in arsenic resistance are now being
gradually translated into strategies for engineering
microbes for effective arsenic remediation. Although
the initial reports are promising, substantial improve-
ments are necessary to move these approaches from the
bench to practice. In this respect, new tools in synthetic
biology will certainly enable us to increase our efforts
toward this end.
AcknowledgementsThe financial support from NSF and U.S. EPA are gratefully acknowledged.
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29.�
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An important paper describing arsenate reductase and showing itsimportance for arsenic resistance.
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46.��
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A concise review about arsenic metabolism in plants and how geneticengineering can improve arsenic phytoremediation. This part is notdetailed in our review.
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47. Tripathi RD, Srivastava S, Mishra S, Singh N, Tuli R, Gupta DK,Maathuis FJM: Arsenic hazards: strategies for tolerance andremediation by plants. Trends Biotechnol 2007, 25:158-165.
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53. Liu J, Liu YP, Powell DA, Waalkes MP, Klaassen CD: Multidrug-resistance mdr1a/1b double knockout mice are moresensitive than wild type mice to acute arsenic toxicity, withhigher arsenic accumulation in tissues. Toxicology 2002,170:55-62.
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55.��
Liu ZJ, Boles E, Rosen BP: Arsenic trioxide uptake by hexosepermeases in Saccharomyces cerevisiae. J Biol Chem 2004,279:17312-17318.
This paper clearly demonstrated that hexose permeases catalyze themajority of the transport of arsenite in S. cerevisiae.
56. Liu ZJ, Sanchez MA, Jiang X, Boles E, Landfear SM, Rosen BP:Mammalian glucose permease GLUT1 facilitates transport ofarsenic trioxide and methylarsenous acid. Biochem BiophysRes Commun 2006, 351:424-430.
57.�
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This paper reported the two major pathways for arsenic detoxification inS. cerevisiae. These results clearly demonstrated that Arr3p and Ycf1prepresent separated pathways for the detoxification of arsenite in yeast.
58. Mukhopadhyay R, Shi J, Rosen BP: Purification andcharacterization of Acr2p, the Saccharomyces cerevisiaearsenate reductase. J Biol Chem 2000, 275:21149-21157.
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This study showed that the vacuolar serine carboxypeptidases CPY andCPC are responsible for PC synthesis in S. cerevisiae.
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74. Singh S, Lee W, DaSilva NA, Mulchandani A, Chen W: Enhancedarsenic accumulation by engineered yeast cells expressingArabidopsis thaliana phytochelatin synthase. BiotechnolBioeng 2008, 99:333-340.
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80. Dameron CT, Winge DR: Peptide-mediated formation ofquantum semiconductors. Trends Biotechnol 1990, 8:3-6.
81. Krumov N, Oder S, Perner-Nochta I, Angelov A, Posten C:Accumulation of CdS nanoparticles by yeasts in a fed-batchbioprocess. J Biotechnol 2007, 132:481-486.
82. Bobrowicz P, Wysocki R, Owsianik G, Goffeau A, Ulaszewski S:Isolation of three contiguous genes, ACR1, ACR2 and ACR3,involved in resistance to arsenic compounds in the yeastSaccharomyces cerevisiae. Yeast 1997, 13:819-828.
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85. Li Y, Cockburn W, Kilpatrick J, Whitelam GC: Cytoplasmicexpression of a soluble synthetic mammalian metallothionein-alpha domain in Escherichia coli — enhanced tolerance andaccumulation of cadmium. Mol Biotechnol 2000, 16:211-219.
86. Bae W, Chen W, Mulchandani A, Mehra RK: Enhancedbioaccumulation of heavy metals by bacterial cells displayingsynthetic phytochelatins. Biotechnol Bioeng 2000, 70:518-524.
87. Bae W, Mehra RK, Mulchandani A, Chen W: Genetic engineeringof Escherichia coli for enhanced uptake and bioaccumulationof mercury. Appl Environ Microbiol 2001, 67:5335-5338.
88.��
Dhankher OP, Li YJ, Rosen BP, Shi J, Salt D, Senecoff JF,Sashti NA, Meagher RB: Engineering tolerance andhyperaccumulation of arsenic in plants by combining arsenatereductase and gamma-glutamylcysteine synthetaseexpression. Nat Biotechnol 2002, 20:1140-1145.
An excellent paper showing how the arsenic defense mechanism inmicrobes can be applied to plants.
89. Song WY, Sohn EJ, Martinoia E, Lee YJ, Yang YY, Jasinski M,Forestier C, Hwang I, Lee Y: Engineering tolerance andaccumulation of lead and cadmium in transgenic plants. NatBiotechnol 2003, 21:914-919.
90.�
Sauge-Merle S, Cuine S, Carrier P, Lecomte-Pradines C, Luu DT,Peltier G: Enhanced toxic metal accumulation in engineeredbacterial cells expressing Arabidopsis thaliana phytochelatinsynthase. Appl Environ Microbiol 2003, 69:490-494.
This paper reported the use of phytochelatin producing E. coli as apotential arsenic accumulating biosorbent.
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92. Wang CL, Maratukulam PD, Lum AM, Clark DS, Keasling JD:Metabolic engineering of an aerobic sulfate reduction
pathway and its application to precipitation of cadmium on thecell surface. Appl Environ Microbiol 2000, 66:4497-4502.
93. Kostal J, Yang R, Wu CH, Mulchandani A, Chen W: Enhancedarsenic accumulation in engineered bacterial cells expressingArsR. Appl Environ Microbiol 2004, 70:4582-4587.
94.��
Singh S, Mulchandani A, Chen W: Highly selective and rapidarsenic removal by metabolically engineered Escherichia colicells expressing Fucus vesiculosus metallothionein. ApplEnviron Microbiol 2008, 74:2924-2927.
For the first time, a metallothionein able to bind to arsenic was utilizedalong with an arsenic transporter for the selective removal of arsenic.Resting cells can be used to completely remove 35 ppb of As(III) in20 min, making this a low-cost option for arsenic removal from water.
95. Dai MH, Copley SD: Genome shuffling improves degradation ofthe anthropogenic pesticide pentachlorophenol bySphingobium chlorophenolicum ATCC 39723. Appl EnvironMicrobiol 2004, 70:2391-2397.
96.��
Crameri A, Dawes G, Rodriguez E, Silver S, Stemmer WPC:Molecular evolution of an arsenate detoxification pathwayDNA shuffling. Nat Biotechnol 1997, 15:436-438.
One of the few reports to show application of irrational approaches forfunctional evolution of the arsenic resistance operon.
97. Chauhan NS, Ranjan R, Purohit HJ, Kalia VC, Sharma R:Identification of genes conferring arsenic resistanceto Escherichia coli from an effluent treatment plantsludge metagenomic library. FEMS Microbiol Ecol 2009,67:130-139.
98. Singh S, Kang SH, Lee W, Mulchandani A, Chen W: SystematicEngineering of Phytochelatin Synthesis and Arsenic Transportfor Enhanced Arsenic Accumulation in E. coli. Biotechnol.Bioeng., in press.
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