Molecular Microbiology - 2004 - M Ller - Redox and Antioxidant Systems of The Malaria Parasite Plasmodium Falciparum

Blackwell Science, LtdOxford, UKMMIMolecular Microbiology0950-382XBlackwell Publishing Ltd, 2004? 200453512911305Review ArticleAntioxidant defence of PlasmodiumS.
Müller
Molecular Microbiology (2004) 53(5), 1291–1305 doi:10.1111/j.1365-2958.2004.04257.x
MicroReview
Redox and antioxidant systems of the malaria parasite

Plasmodium falciparum
Sylke Müller*† superoxide anions, hydrogen peroxide and hydroxyl radi-

School of Life Sciences, Wellcome Trust Biocentre, cals that damage nucleic acids, proteins, lipids and
University of Dundee, UK. membranes and yet allows essential metabolic functions
(Imlay, 2003). In order to preserve this crucial reducing
milieu, aerobes have evolved efficient redox and antioxi-
Summary
dant systems (Sies, 1993; 1997). The redox state of a
The malaria parasite Plasmodium falciparum is highly cell is mediated by the ratios of reduced and oxidized
adapted to cope with the oxidative stress to which it pyridine nucleotides [NAD(P)H/NAD(P)+] and thiols
is exposed during the erythrocytic stages of its life such as glutathione/glutathione disulphide (GSH/GSSG)
cycle. This includes the defence against oxidative and thioredoxin/thioredoxin disulphide [Trx(SH)2/Trx(S)2].
insults arising from the parasite’s metabolism of hae- These ratios are intimately related to the cellular levels
moglobin which results in the formation of reactive of ROS as the activities of many enzymes involved in
oxygen species and the release of toxic ferriprotopor- antioxidant functions depend on the redox state of the
phyrin IX. Central to the parasite’s defences are pyridine nucleotide pools and on the redox state of cel-
superoxide dismutases and thioredoxin-dependent lular thiols such as glutathione and thioredoxin (Bunik,
peroxidases; however, they lack catalase and glu- 2003).
tathione peroxidases. The vital importance of the The intracellular levels of ROS are kept at a low level
thioredoxin redox cycle (comprising NADPH, thiore- by a number of enzymatic and non-enzymatic antioxi-
doxin reductase and thioredoxin) is emphasized by dants. The first lines of defence are superoxide dismu-
the confirmation that thioredoxin reductase is essen- tases (SODs), a family of metalloproteins catalysing the
tial for the survival of intraerythrocytic P. falciparum. dismutation of superoxide anions to form hydrogen per-
The parasites also contain a fully functional glu- oxide and oxygen (Fridovich, 1995). Hydrogen peroxide
tathione redox system and the low-molecular-weight itself needs to be reduced to water and oxygen to pre-
thiol glutathione is not only an important intracellular vent the oxidation of other cellular components. This
thiol redox buffer but also a cofactor for several redox reaction is catalysed by a variety of peroxidases includ-
active enzymes such as glutathione S-transferase and ing the haem-peroxidase catalase, glutathione-depen-
glutaredoxin. Recent findings have shown that in dent peroxidases (GSH-Px) and thioredoxin-dependent
addition to these cytosolic redox systems the parasite peroxidases or peroxiredoxins (Trx-Px) (Sies, 1997; Hof-
also has an important mitochondrial antioxidant mann et al., 2002; Wood et al., 2003). GSH-Px and Trx-
defence system and it is suggested that lipoic acid Px obtain their reducing equivalents from two distinct
plays a pivotal part in defending the organelle from redox systems, the glutathione and the thioredoxin redox
oxidative damage. systems (Arner and Holmgren, 2000; Becker et al.,
2000; Yodoi et al., 2001; Filomeni et al., 2002). Both
comprise a cascade of redox-active proteins and pep-
Introduction
tides that transfer reducing equivalents from NADPH to
Aerobic life requires the maintenance of an appropriate acceptor molecules, in this case hydroperoxides (Fig. 1).
intracellular redox environment that minimizes the gen- Apart from these enzymatic antioxidants, each cell con-
eration of reactive oxygen species (ROS) such as tains a number of non-enzymatic antioxidants such as
tocopherol (vitamin E), ascorbate (vitamin C) and the
Accepted 9 June, 2004. *For correspondence. E-mail tripeptide glutathione, which not only acts as a cofactor
s.muller@dundee.ac.uk; Tel. (+44) 1382 345760; Fax (+44) 1382 for a number of enzymatic reactions but also reduces
345764. †Present address: Division of Biological Chemistry and
Molecular Microbiology, School of Life Sciences, University of ROS non-enzymatically and acts as a reductant for both
Dundee, MSI/WTB complex, Dundee DD1 5EH, Scotland, UK. vitamins (Di Mascio et al., 1991).
© 2004 Blackwell Publishing Ltd

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1292 S. Müller
means to prevent membrane damage and parasite death
(Atamna and Ginsburg, 1993; Tilley et al., 2001; Becker
et al., 2004) (Fig. 2).
Notably, not only is the parasite itself under oxidative
stress but the host cell also shows oxidative alterations
when infected with Plasmodium. This is demonstrated by
changes in erythrocyte membrane fluidity, most probably
because of alterations of erythrocyte membrane lipid com-
position and protein cross-linking (Destro Bisol, 1999; Gir-
ibaldi et al., 2001; Williams et al., 2002; Omodeo-Sale
et al., 2003). Haemichrome accumulation on the inner
surface of the parasitized erythrocytes as well as the
aggregation of erythrocyte band 3 and the increasing
occurrence of auto-anti-band 3 antibodies suggest that
Fig. 1. Thioredoxin and glutathione redox systems. The thioredoxin the host erythrocytes are severely oxidatively damaged by
and glutathione redox cycles comprise a number of redox active
proteins that transfer reducing equivalents from NADPH to an accep-
the Plasmodium infection (Parker et al., 2004). This is
tor molecule. Both systems have a range of common acceptors such most apparent in erythrocytes infected with late parasite
as ribonucleotide reducase and transcription factors. In addition, stages (Giribaldi et al., 2001). These modifications are
thioredoxin reduces peroxiredoxins which are not substrates of the
glutathione redox cycle. The thioredoxin and glutathione redox cycles
reminiscent of those found in erythrocytes of humans with
in Plasmodium seem to be linked by the reduction of GSSG via red blood cell disorders such as sickle cell anaemia and
thioredoxin, a situation that may be of physiological relevance under a- and b-thalassaemias and also glucose 6-phosphate
enhanced oxidative stress (Kanzok et al., 2000). Glutaredoxin effi-
ciently reduces plasmoredoxin, which is only a very poor substrate
dehydrogenase deficiency and it is believed that the
for thioredoxin. Thus, the two redox systems have overlapping func- underlying mechanism for these transformations is the
tions but also play unique roles that very probably make both systems enhanced oxidative stress in the defective erythrocytes
indispensable for P. falciparum.
TrxR-ox, oxidized form of thioredoxin reductase; TrxR-red, reduced
(Shalev et al., 1995; Beppu et al., 1996; Shalev and Heb-
form of thioredoxin reductase; Trx-ox, oxidized from of thioredoxin; Trx- bel, 1996; de Jong et al., 1997; Aslan et al., 2000). Inter-
red, reduced from of thioredoxin; GR-ox, oxidized form of glutathione estingly, these red blood cell disorders and also glucose
reductase; GR-red, reduced form of glutathione reductase; GSH,
reduced form of glutathione; GSSG, glutathione disulphide; Grx-ox,
6-phosphate dehydrogenase deficiency confer a certain
oxidized form of glutaredoxin; Grx-red, reduced form of glutaredoxin. degree of resistance to an infection with Plasmodium and
often limit the severity of the disease (Roberts and Will-
iams, 2003). One hypothesis to explain this is that the
Oxidative stress in Plasmodium falciparum-infected
increased oxidative stress within the defective erythrocyte
erythrocytes
causes an impaired infection and growth rate of the par-
It has been shown that malaria parasites are particularly asites (Eaton et al., 1976; Friedman, 1978; Pasvol et al.,
vulnerable to oxidative stress during their erythrocytic life 1978). Another hypothesis (Cappadoro et al., 1998; Gir-
stages (Hunt and Stocker, 1990; Simoes et al., 1992; ibaldi et al., 2001; Williams et al., 2002) suggests that
Müller et al., 2003; Becker et al., 2004). This is not sur- defective erythrocytes infected with early stages of P. fal-
prising as the parasites live in a pro-oxidant environment ciparum are more efficiently phagocytized by the host’s
that contains oxygen and iron, the key prerequisite for the immune system because of the earlier occurrence of band
formation of ROS via the Fenton reaction (see Fig. 2). 3 cross-linking which results in the early appearance of
Haemoglobin is taken up by the parasites into their acid band 3 auto-antibodies. Thus, the parasitized erythrocytes
food vacuole which leads to the spontaneous oxidation of are recognized by the host’s immune system at an early
Fe2+ to Fe3+ and the formation of superoxide anions. This stage of the infection and the parasitaemia of the infected
combination inevitably leads to the generation of hydrogen individuals is kept low. Similar changes also occur in nor-
peroxide and subsequently hydroxyl radicals, both highly mal Plasmodium-infected erythrocytes but at a much later
reactive and toxic oxygen intermediates (Fig. 2) (Liochev stage of the infection when most Plasmodium-infected
and Fridovich, 1999). Furthermore, toxic haem (ferri/ cells are already sequestered in the host’s capillary sys-
ferroprotoporphyrin IX; FP IX) is released upon haemo- tem and so no longer circulating such that they are not
globin digestion and this must be detoxified. Most of the recognized by the host’s immune system as efficiently.
released FP IX is biomineralized (up to 90%; Egan et al., Much less is known about other developmental stages
2002) to form inert haemozoin but there are reports that of Plasmodium as they are much less amenable for
an appreciable amount of FP IX (even as much as 50%; detailed physiological investigations but it has been sug-
Loria et al., 1999; Zhang et al., 1999) escapes biominer- gested that the mosquito stages, especially invading ook-
alization and has to be degraded or sequestered by other inetes and sporozoites, are exposed to elevated oxidative
© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

Antioxidant defence of Plasmodium 1293
Fig. 2. Sources of reactive oxygen species in Plasmodium falciparum. The major source for reactive oxygen species in P. falciparum during its
intraerythrocytic life is the digestion of host cell haemoglobin in the parasite’s food vacuole. Free haem (FP IX) is released from the digested
haemoglobin and the majority (up to 90%; Egan et al., 2002) is biomineralized to haemozoin. However, some free FP IX (up to 50% according
to Loria et al., 1999; Zhang et al., 1999) is released from the food vacuole into the parasite cytosol where it causes membrane damage and can
undergo redox reactions which lead to the generation of superoxide anions in this parasite compartment. Superoxide anions, resulting from the
oxidation of haem-iron in haemoglobin, are either detoxified by SOD, to yield H2O2, or can react in a spontaneous reaction with H2O2 leading to
the formation of hydroxyl radicals. In addition, hydroxyl radicals are generated in the presence of free iron via the Fenton reaction. These radicals
are highly reactive and cause, for instance, lipid peroxidation. H2O2 generated by the SOD reaction has to be detoxified by reduction to H2O. In
P. falciparum, this is exclusively achieved by thioredoxin peroxidases because the parasites lack catalase and glutathione peroxidases.
FP IX, ferri/ferroprotoporphyrin IX; HbO-Fe2+, oxy-haemoglobin containing ferriprotoporphyrin IX; MetHb-Fe3+, methaemoglobin containing ferro-
protoporhpyrin IX; SOD, superoxide dismutase.
and nitrosative stress elicited by the mosquito’s innate gets for a variety of infectious diseases (Turrens, 2004).
immune response and that this causes major parasite However, the nature of the substrate, a very small mole-
losses (Han et al., 2000; Dimopoulos et al., 2002; Kumar cule, has made it extremely difficult to identify substrate-
et al., 2003). Nevertheless, some ookinetes manage to or product-analogues that would be suitable parasite-spe-
manifest an infection within the mosquito’s midgut and cific inhibitors for these enzymes.
infective sporozoites still reach the vector’s salivary glands Plasmodium falciparum possesses two distinct genes
and it seems likely that these parasite stages also rely on that encode different SODs. One of the proteins is a
their antioxidant systems to prevent damage caused by cytosolic, Fe-dependent SOD, called SOD-1 that appears
ROS and reactive nitrogen species. However, the studies to be transcribed and expressed throughout the erythro-
to date are only indicative and more work is required to cytic cycle of the parasite (Gratepanche et al., 2002; Sien-
analyse the roles of the parasite’s antioxidant proteins kiewicz et al., 2004). This protein was also described from
facilitating a successful infection of the mosquito host. P. ovale, P. malariae and P. vivax as well as in the related
apicomplexan parasite Toxoplasma gondii (Dive et al.
2003). As SOD-1 is a cytosolic protein, it is unlikely that
Redox and antioxidant systems in P. falciparum it acts on superoxide anions produced during haemoglo-
erythrocytic stages bin digestion in the parasite’s food vacuole. Thus, it still
remains a possibility that the large amount of Cu/Zn-SOD
Superoxide dismutases
taken up by the parasite from the host erythrocyte con-
Superoxide dismutases are metalloproteins that use tributes to the detoxification of superoxide anions in this
copper/zinc (Cu/Zn), iron (Fe) or manganese (Mn) as organelle (Fairfield et al., 1983). Furthermore, it cannot be
metal cofactors. Eukaryotes possess cytosolic Cu/Zn- excluded that a large proportion of the superoxide anions
dependent SOD and mitochondrial Mn-dependent SODs, generated in the food vacuole spontaneously dismutate
whereas protozoan parasites generally appear to contain because of the acidic pH in this parasite organelle. The
only Fe-dependent SODs (Fridovich, 1995). This poten- second P. falciparum SOD, named SOD-2, is a mitochon-
tially renders the parasite SODs attractive therapeutic tar- drial protein. Notably the long N-terminal extension of this

1294 S. Müller
protein is reminiscent of a bipartite apicoplast targeting buffer in almost all aerobic cells (Sies, 1999). The ratio
sequence (Foth et al., 2003) which prompted the predic- between GSH and GSSG is usually between 10:1 and
tion that it is an apicoplast-localized protein. However, 100:1 – it is maintained far on the side of the reduced form
overexpressing a SOD-2–green fluorescent protein (GFP) of glutathione. This is mainly achieved by the action of
fusion in P. falciparum revealed that the enzyme’s target- glutathione reductase (GR) but there also exist GSSG-
ing sequence conferred transport of GFP into the para- efflux pumps that export excess GSSG in order to main-
site’s mitochondrion (Sienkiewicz et al., 2004). This tain an adequate intracellular redox balance (Krauth-Sie-
highlights the dangers of relying too heavily on predictions gel et al., 1989; Oude et al., 1990; Homolya et al., 2003).
from gene sequences. Interestingly, one of the SODs of It has been shown that the reversible formation of protein-
the related apicomplexan parasite T. gondii possesses a GS-mixed disulphides also aids to maintain an adequate
similar untypical mitochondrial targeting sequence, sug- GSH/GSSG ratio (Thomas et al., 1995; Sies, 1999). In
gesting that apicomplexan parasites might have unusual addition to these reactions, the de novo synthesis of the
and distinct mechanisms that confer transport into this tripeptide also contributes to sustaining sufficiently high
organelle (Brydges and Carruthers, 2003). The presence intracellular GSH levels (Griffith, 1999). Apart from its role
of a mitochondrial SOD seems imperative in Plasmodium as a general thiol redox buffer, GSH acts as a cofactor for
as the parasites clearly possess an active respiratory a variety of proteins including glutathione-dependent per-
chain (Kita et al., 2002) which unavoidably leaks superox- oxidases, glutathione S-transferases (GSTs), glutaredox-
ide anions. These have to be detoxified by a mitochondrial ins and glyoxalases (Sies, 1999). It is also directly involved
SOD to prevent damage to the metabolic functions, in antioxidant reactions – for instance, the termination of
nucleic acids, proteins and membranes of the organelle radical-based chain reactions where single electrons are
(Inoue et al., 2003). transferred from thiyl radicals or disulphide radicals (Frey,
So far, no apparent apicoplast SOD gene has been 1997).
identified in the genome of P. falciparum, possibly implying The glutathione metabolism of Plasmodium-infected
that the organelle does not require this enzymatic activity erythrocytes has been the focus of a number of studies
because little or no superoxide anions are generated by which have revealed that the roles of the tripeptide are
the metabolic reactions present in the organelle. quite numerous and not only limited to its redox and anti-
oxidant functions (Fig. 3) (Becker et al., 2003a). Notably,
it has been suggested that GSH is involved in drug resis-
Control of intracellular glutathione levels in P. falciparum
tance both as a cofactor for enzymatic reactions (e.g.
The tripeptide glutathione (GSH; g-glutamyl-cysteinyl- GST) and helping to mediate resistance as a source of
glycine) is the major low-molecular-weight thiol redox reductive detoxification of FP IX, the toxic by-product of
Fig. 3. Control of intracellular glutathione

levels in Plasmodium falciparum. The major
low-molecular-weight thiol in P. falciparum is
glutathione. It is imperative that an appropriate
ratio between reduced glutathione (GSH) and
glutathione disulphide (GSSG) is maintained
and a number of reactions have been identified
in the parasite that contribute to sustaining this
crucial redox balance. GSSG formed during the
non-enzymatic degradation of FP IX or the
reduction of glutaredoxin (Grx) is reduced effi-
ciently by glutathione reductase (GR) back to
GSH. In addition, excess GSSG is effluxed from
the parasite by a GSSG-efflux system. In order
to refill the GSH-pool, the tripeptide is synthe-
sized de novo by g-glutamylcysteine syn-
thestase (g-GCS) and glutathione synthetase
(GS). In addition, P. falciparum possesses a
single glutathione S-transferase (GST) that acts
as a detoxifying enzyme binding electrophilic
harmful compounds (X) to form GS-X com-
plexes that are very probably effluxed from the
parasite by a GS-X pump. GST also acts as an
alky hydroperoxide (ROOH) peroxidase. The
GSSG that arises from the reductive reactions
of GST can enter the glutathione redox cycle
via GR and be recycled back to GSH.
FP IX, ferri/ferroprotoporphyrine IX; g-GluCys,
g-glutamylcysteine.

haemoglobin digestion, through which chloroquine exerts have deleterious effects for the parasites (Atamna et al.,
its action (see Glutathione-dependent detoxification sys- 1996).
tems in P. falciparum) (Dubois et al., 1995; Ginsburg et al.,
1998; Harwaldt et al., 2002; Liebau et al., 2002). Glutathione synthesis. Plasmodium parasites possess a
functional glutathione synthesis pathway – the genes for
Glutathione reductase. Glutathione reductase is the g-glutamylcysteine synthetase (g-GCS) and glutathione
enzyme responsible for keeping glutathione in its reduced synthetase (GS), the two enzymes that catalyse the liga-
state (Fig. 3). GR is a member of a group of enzymes tion of glutamate and cysteine followed by the addition of
known as flavo disulphide oxidoreductases which also glycine, have been isolated and characterized (Fig. 3)
contains thioredoxin reductase, mercuric ion reductase (Lüersen et al., 1999; 2000; Meierjohann et al., 2002b).
and trypanothione reductase (Williams, 1992). Plasmo- The g-GCS genes of different P. falciparum strains contain
dium GR has attracted a lot of attention as a potential a variable number of repeats in the N-terminal region,
therapeutic target against malaria (Schirmer et al., 1995; which do not occur in the P. berghei g-GCS gene (Birago
Färber et al., 1996; 1998; Gilberger et al., 2000) and it is et al., 1999; Lüersen et al., 1999). These variations
hoped that the knowledge of the three-dimensional struc- appear to be linked neither with drug resistance nor with
tures of human and Plasmodium GRs (Karplus and geographical distribution of the parasites and their role
Schulz, 1987; Sarma et al., 2003) will aid in the endeav- (if any) is not yet understood (Lüersen et al., 1999).
ours to design rationally inhibitors that specifically target Plasmodium-infected erythrocytes are killed by the spe-
the parasite protein. The primary structure of Plasmodium cific g-GCS inhibitor D,L-buthionine-S,R-sulphoximine
GR contains parasite-specific insertions in the FAD- (BSO) because of the depletion of glutathione from the
domain (residues 123–134), the central domain (residues Plasmodium-infected cell (Lüersen et al., 2000). Plasmo-
314–347) and the interface domain (residues 496–499). dium-infected erythrocytes rapidly lose glutathione via a
The first two insertions have been experimentally deleted GSSG efflux pump (Atamna and Ginsburg, 1997; Ayi
and this affected the stability and FAD cofactor binding et al., 1998; Ginsburg et al., 1998; Lüersen et al., 2000;
capacity, respectively, of the protein (Gilberger et al., Meierjohann et al., 2002a). When GSH de novo synthesis
2000). Unfortunately, the structural analyses of Plasmo- is inhibited, half of the cellular glutathione is lost within
dium GR so far reported did not resolve the structures of 2 h, suggesting that the parasitized cell is highly depen-
the insertions, thus it is presumed that they are flexible dent on the functional synthesis of the tripeptide. Conse-
loops that are most probably located on the protein’s quently, inhibition of GSH synthesis is lethal for the
surface. Therefore, it is believed that they only marginally parasites presumably because GSH pools decrease dra-
affect the overall structure of the protein (Sarma et al., matically and the GSH/GSSG ratio cannot be maintained
2003). Overall, the structures of Plasmodium GR and (Atamna and Ginsburg, 1997; Ayi et al., 1998; Lüersen
human GR are very similar. However, the subtle differ- et al., 2000; Meierjohann et al., 2002a). In spite of an
ences in the cavity of the dimer interface may allow the active GSH de novo synthesis and a functional glutathione
design of inhibitors that specifically bind in this area of the redox system, Plasmodium-infected erythrocytes contain
parasite protein (Sarma et al., 2003; Becker et al., 2004). only about half the GSH concentration of non-infected
Indeed, it has been suggested that tricyclic heteroaro- erythrocytes (Lüersen et al., 2000; Meierjohann et al.,
matic compounds such as isoalloxazine derivatives and 2002a), which is possibly the result of the high efflux rate
methylene blue which inhibit the enzymatic activity with Ki of GSSG observed in infected erythrocytes (Fig. 3). Inter-
values in the low micromolar range exert their effects estingly, Ginsburg and colleagues (Atamna and Ginsburg,
through this area of the protein (Färber et al., 1998; 1997) suggested that the GSSG effluxed from Plasmo-
Sarma et al., 2003; Becker et al., 2004). Notably, methyl- dium might be a source of glutathione for the infected host
ene blue was the first synthetic compound used in clinical erythrocytes (which contain high levels of GR) and aids
anti-malarial therapy and its anti-malarial activity was to maintain the reducing environment of the host cell and
reported by Guttmann and Ehrlich (1891). Although Plas- so supports parasite development. However, our subse-
modium GR might be one of the molecular targets of quent results suggest that GSSG is actually entirely lost
methylene blue, it is unlikely to be the primary one as the from the Plasmodium-infected cell as effluxed glutathione
IC50 of parasite growth inhibition by methylene blue is in is found in the culture medium rather than the erythrocyte
the low nanomolar range and thus one order of magnitude compartment (Lüersen et al., 2000; Meierjohann et al.,
lower than the Ki for GR inhibition by methylene blue 2002a). This implies that despite the presence of efficient
(Meierjohann et al., 2002a). This strongly suggests that parasite and host cell GRs, GSSG cannot be reduced at
the compound has additional effects that lead to parasite sufficient rates to prevent GSSG efflux from the Plasmo-
death (Sarma et al., 2003). In fact, it has been shown that dium-infected cell. This furthermore suggests that the
it inhibits the formation of hemozoin which obviously can increased oxidative stress generated by the infection with

1296 S. Müller
Plasmodium causes the oxidation of such large amounts appear to have developed between chloroquine-sensitive
of GSH that there needs to be an increased GSSG-efflux and chloroquine-resistant Plasmodium species (Ursos
to maintain an adequate GSH/GSSG thiol redox state in and Roepe, 2002) including changes in their glutathione
the infected cell. Interestingly, glucose 6-phosphate dehy- metabolism (Meierjohann et al., 2002a; Perez-Rosado
drogenase-deficient erythrocytes also have decreased et al., 2002). Using BSO as a research tool to deplete
GSH levels, which might be explained by the decreased GSH levels in Plasmodium-infected erythrocytes, it was
availability of NADPH for the reduction of the tripeptide shown that the efficacy of chloroquine in P. falciparum and
and so its efflux from the infected cell (Cappadoro et al., P. berghei strains resistant to the drug was increased
1998). These observations together suggest that one (Dubois et al., 1995; Ginsburg et al., 1998; Famin et al.,
reason for the increased oxidative damage observed in 1999; Platel et al., 1999). The data suggest that there is
Plasmodium-infected erythrocytes and also glucose 6- indeed a causal correlation between intracellular GSH
phosphate dehydrogenase-deficient erythrocytes may be levels and chloroquine action.
the inadequate concentration of intracellular glutathione. In addition to non-enzymatic detoxification of FP IX,
As BSO acts specifically on g-GCS, it can be used as a other mechanisms of FP IX removal have been proposed
research tool that efficiently depletes GSH without affect- and these include its sequestration by parasite proteins.
ing other metabolic pathways or proteins. However, the Proteins possibly involved in this process include histi-
inhibitor not only acts on the parasite enzyme but also dine-rich protein 2 (Choi et al., 1999), glyceraldehyde-3-
inhibits the host cell g-GCS activity (Meierjohann et al., phosphate dehydrogenase, protein disulphide isomerase,
2002a). Therefore it remains to be shown that it is indeed GR (Campanale et al., 2003) and GST (Harwaldt et al.,
the inhibition of Plasmodium g-GCS that impairs parasite 2002; Liebau et al., 2002). Apart from histidine-rich pro-
survival and knockout studies of the Plasmodium gene are tein 2, which in complex with FP IX appears to develop
underway to address this point. peroxidase-like activity (Mashima et al., 2002), most of the
other FP IX binding proteins are in fact inhibited by FP IX.
Therefore, it is difficult to judge whether their affinity to FP
Glutathione-dependent detoxification systems in
IX is actually a protective mechanism or whether binding
P. falciparum
of FP IX has a deleterious effect on the parasites. In case
Apart from its role as an antioxidant and thiol redox buffer, of glyceraldehyde-3-phosphate dehydrogenase, it has
glutathione is involved in a variety of vital detoxification been suggested that the strong inhibition by FP IX is a
reactions in P. falciparum. Despite the efficient biominer- mechanism by which the parasite is adapted to survive
alization of free haem into haemozoin upon haemoglobin FP IX stress. The inhibition of glyceraldehdye-3-
digestion by the parasite, a considerable amount of toxic phosphate dehydrogenase under elevated FP IX stress
FP IX remains free (Loria et al., 1999; Zhang et al., 1999). results in glucose being primarily metabolized via the hex-
Free FP IX is toxic because it has detergent-like proper- ose monophosphate shunt rather than glycolysis and this
ties interfering with membrane integrity and has the ability provides increased levels of NADPH that are required for
to undergo redox reactions causing the generation of ROS antioxidant reactions in the parasite cell (Campanale
because of the bound iron. Therefore, it needs to be et al., 2003).
detoxified or sequestered to prevent membrane toxicity An enzyme that is directly dependent on the availability
and ultimately parasite death. Ginsburg and colleagues of glutathione is GST. GSTs catalyse the conjugation of
have shown that free FP IX is degraded non-enzymatically GSH to the electrophilic centres of hydrophobic com-
by GSH and that this mechanism is inhibited by chloro- pounds and thereby detoxify a wide range of mutagens,
quine (Atamna and Ginsburg, 1995; Zhang et al., 1999; carginogenic, pharmacological active molecules and by-
Deharo et al., 2003). Thus a depletion of GSH would products of oxidative stress. Unlike higher eukaryotes,
result in a less efficient detoxification of free FP IX and Plasmodium only possesses a single gene encoding GST.
consequently parasite death. The resistance of P. falci- Interestingly, other parasitic protozoa such as Leishmania
parum to chloroquine is one of the major drawbacks in the and Trypanosoma cruzi also contain few GSTs and have
fight against severe malaria, as this drug has been highly either unusual glutathione-dependent S-transferases or
efficient, has no side-effects and is affordable. A number S-transferase activities that depend on their unique thiol
of different mechanisms by which the parasites have trypanothione (Ouaissi et al., 1995; Denton et al., 2004;
developed resistance to chloroquine have been sug- Vickers and Fairlamb, 2004). This low number of GSTs
gested and mutations of the P. falciparum chloroquine possibly reflects that parasitic protozoa do not require a
resistance transporter Pfcrt and the P-glycoprotein Phg-1 great number of xenobiotic detoxification enzymes and
are the most favoured theories at the moment (Fidock that GSTs in parasitic protozoa have quite different func-
et al., 2000; Reed et al., 2000; Mu et al., 2003; Tran and tions from those found in other eukaryotes. Two potential
Saier, 2004). However, many other metabolic differences physiological functions of the P. falciparum have been

proposed. (i) In vitro the enzyme acts as an alkyl hydro- the thioredoxin superfamily which are characterized by a
peroxide peroxidase which might have an essential role specific active site CXXC motif (Table 1) (Rahlfs et al.,
for parasite survival as glutathione peroxidases, enzymes 2003). Despite the low degree of similarity between their
that reduce these metabolic by-products in other eukary- primary sequences, the proteins are structurally related –
otes, are absent from P. falciparum. (ii) The protein might they all possess the so-called thioredoxin fold and
mediate the efficient sequestration of free FP IX through have similar functions within the cell (Holmgren, 2000;
binding to it, an act that results in severe inhibition of the Fernandes and Holmgren, 2004). They are considered
GST (Harwaldt et al., 2002; Liebau et al., 2002). A number redox messengers that interact with a variety of redox
of questions arise from this latter hypothesis, e.g. what active proteins and metabolites such as ribonucleotide
happens to GST which has FP IX bound – is it degraded reductase, transcription factors, peroxidases, cyclophilins
by the parasite? If it is degraded, is FP IX released again? and low-molecular-weight thiols (Holmgren, 2000; Moto-
Are GST–FP IX complexes sequestered in the parasite or hashi et al., 2003; Fernandes and Holmgren, 2004).
are they exported from the parasite into the host cell Plasmodium falciparum possesses one classical glu-
cytosol? These important aspects remain to be clarified. taredoxin with the typical CPYC redox active site (Rahlfs
The three-dimensional structure of Plasmodium GST et al., 2001). This active site motif allows glutaredoxin to
shows some similarities to Mu-class GSTs (Fritz-Wolf interact with proteins containing sulphydryl groups that
et al., 2003; Perbandt et al., 2004). However, the parasite need to be either oxidized or reduced, depending on their
GST appears to have a larger hydrophobic binding pocket distribution, the redox state of the cell and their abun-
than the Mu-class mammalian GSTs, which is also more dance. Plasmodium glutaredoxin was shown to reduce
solvent-accessible because the C-terminus of the protein ribonucleotide reductase and it acts as an efficient reduc-
is truncated in comparison to other GSTs. Usually the C- tant for plasmoredoxin, another member of the thioredoxin
terminal part of Mu-class GSTs structurally restricts entry superfamily that appears to be unique to Plasmodium
of substrates into the hydrophobic binding pocket and thus parasites (Rahlfs et al., 2001; 2003; Becker et al., 2003b).
it is believed that the substrate specificity of P. falciparum Plasmoredoxin itself also reduces ribonucleotide reduc-
GST is less restricted which allows for the detoxification tase but its precise function in Plasmodium remains to be
of a wider range of molecules. This could also explain why elucidated. In addition, Plasmodium possesses a genuine
these parasites do not require as many GSTs as do other thioredoxin (Kanzok et al., 2000; Krnajski et al., 2001a)
eukaryotes. It also has been suggested that this structural which is reduced by thioredoxin reductase and confers
difference might be exploitable for the design of parasite- reduction of the 2-Cys containing peroxiredoxins (Krnajski
specific inhibitors acting against the enzyme and some et al., 2001b; Rahlfs et al., 2001; Akerman and Müller,
peptide-based inhibitors have already been tested, 2003), ribonucleotide reductase (Rahlfs et al., 2003) and
although without any success (Fritz-Wolf et al., 2003). GSSG (Kanzok et al., 2000). Thus all three members of
Unfortunately, neither of the structural analyses refers to the thioredoxin superfamily have overlapping functions in
the potential binding site for FP IX, so that this intriguing Plasmodium. Other organisms such as bacteria and yeast
phenomenon remains to be investigated in future struc- usually contain more than one glutaredoxin and thiore-
tural studies. doxin that show some functional redundancy (Grant,
2001; Ritz and Beckwith, 2001; Garrido and Grant, 2002).
However, analyses by gene knockout studies in bacteria
Glutaredoxin, thioredoxin and thioredoxin-like proteins in
and yeast have revealed that each of the proteins also
P. falciparum
plays distinct roles in response to changes in growth con-
Plasmodium possess a number of proteins belonging to ditions and oxidative stress (Aslund and Beckwith, 1999;
Table 1. Genes encoding small redox active proteins in P. falciparum.
Gene Protein Location Predicted localizationa
trx 1 Thioredoxin-WCGPC-motif Chr.14 (PF14_0545) Cytosol

tlp-1 Thioredoxin-like protein-1-WCGPC-motif Chr.14 (PF14_0590) Cytosol
tlp-2 Thioredoxin-like protein-2-WCAPC-motif Chr.9 (PFI1250w) Cytosol
tlp-3 Thioredoxin-like protein-3-WCQAC-motif Chr.13 (MAL13P1.225) Unclear
tlp-4 Thioredoxin-like protein-4-WCKPC-motif Chr.9 (PFI0790w) Cytosol
grx-1 Glutaredoxin 1-CPYC-motif Chr.3 (PFC0271c) Cytosol
glp-1 Glutaredoxin-like protein-1-CGFS-motif Chr.3 (MAL3P2.10) Mitochondrial
glp-2 Glutaredoxin-like protein-2-CKFS-motif Chr.6 (MAL6P1.72) Cytosol
plrx Plasmoredoxin-WCKYC-motif Chr.3 (PFC0166w) Cytosol
a. The localization of the proteins was predicted using TargetP (Emanuelsson et al., 2000), PlasMit (Bender et al., 2003) and PlasmoAP (Foth
et al., 2003).

1298 S. Müller
Grant, 2001). Similar studies are still to be performed in design of specific inhibitors against the parasite enzyme.
Plasmodium parasites in order to dissect the precise Importantly, the P. falciparum trxR gene has been genet-
physiological roles that these functionally and structurally ically validated as a potential drug target as its disruption
related proteins have for the maintenance of the intracel- proved to be lethal for the parasites (Krnajski et al., 2002).
lular redox balance and the response to environmental Thus, the protein provides good potential for the design
changes of the parasite. It is, however, very likely that the of inhibitors that specifically target parasite survival. In a
biological functions of thioredoxin are essential for para- first attempt, 350 000 compounds were screened and
site survival as the knockout of thioredoxin reductase has unsaturated Mannich bases were identified as potential
a lethal effect on P. falciparum (Krnajski et al., 2002). In chemical leads (Davioud-Charvet et al., 2003).
addition to the classical thioredoxin and glutaredoxin and
the unusual plasmoredoxin, Plasmodium also possesses
Thioredoxin-dependent peroxidases and peroxiredoxins
glutaredoxin- and thioredoxin-like proteins (Rahlfs et al.,
2001; 2003) which are thought to be involved in the reduc- Peroxiredoxins are a ubiquitous group of peroxidases that
tion of protein-GS-mixed disulphides that are inevitably exert their reductive activity via active-site cysteine resi-
formed during exposure to ROS and generally lead to a dues. This protein family is structurally distinguished into
reversible inactivation of protein functions. three classes – the typical 2-Cys peroxiredoxins, the atyp-
ical 2-Cys peroxiredoxins and the 1-Cys peroxiredoxins
(Wood et al., 2003). The number of active-site cysteines
Thioredoxin reductase
involved in catalysis determines the classification of the
Plasmodium has a functional glutathione/glutaredoxin different family members. Plasmodium possesses three
redox system, but they also contain the proteins compris- genes encoding genuine peroxiredoxins, which is not
ing the thioredoxin redox system – thioredoxin reductase, unusual as most organisms possess more than one per-
thioredoxin and thioredoxin-dependent peroxidases. oxiredoxin (Wood et al., 2003; Wong et al., 2004). Two of
Thioredoxin reductase has been identified in prokaryotes the Plasmodium proteins are typical 2-Cys peroxiredoxins
and eukaryotes and found to exist in two different forms. and one is a member of the 1-Cys peroxiredoxin family
In bacteria, fungi, plants and some protozoan parasites (Krnajski et al., 2001b; Rahlfs and Becker, 2001). One of
such as Trichomonas vaginalis, the protein is 35 kDa; the 2-Cys peroxiredoxins and the 1-Cys peroxiredoxin are
whereas in mammals, insects and Plasmodium falci- cytosolic proteins whereas the second 2-Cys peroxire-
parum, thioredoxin reductase is between 55 and 60 kDa doxin possesses a potential mitochondrial targeting
(Williams et al., 2000; Coombs et al., 2004). Phylogenetic sequence suggesting that the parasites possess both a
analyses revealed that the proteins are evolutionarily functional cytosolic and mitochondrial thioredoxin redox
unrelated and that high-Mr TrxR are more related to GR cycle. Notably the gene encoding the mitochondrial 2-Cys
than to the low-Mr TrxR (Hirt et al., 2002). In addition, the peroxiredoxin is transcribed throughout the erythrocytic
catalytic mechanisms of high- and low-Mr TrxR differ fun- cycle of parasite development but the transcription levels
damentally (as reviewed by Williams et al., 2000) which appear to be higher in gametocytes and sporozoites (Le
should allow the design of specific inhibitors against the Roch et al., 2003). This is consistent with the observation
low-Mr TrxR which have therapeutic potential. that the mitochondrion of these parasite stages is more
The TrxR of Plasmodium is high-Mr and in this way metabolically active which means that a functional mito-
similar to the mammalian enzyme – but there are signifi- chondrial antioxidant defence might be of greater impor-
cant differences between the active sites of parasite and tance to these stages of the parasite.
host proteins (Williams et al., 2000). Similar to GR, high- In addition to these genuine peroxiredoxins, Plasmo-
Mr TrxR are homodimeric, NADPH-dependent disulphide dium possesses a glutathione peroxidase-like protein that
oxidoreductases that possess one FAD cofactor bound preferably uses thioredoxin as a reducing cofactor (Sztajer
per subunit and residues from both subunits contribute to et al., 2001). Notably, the recombinant protein analysed in
the two active sites per homodimer (Gilberger et al., 1997; the study performed by Sztajer et al. (2001) turned out to
Krnajski et al., 2000). The major difference to GR is the be N-terminally truncated by the bacterial expression sys-
presence of a second pair of redox-active residues in tem – suggesting that the protein’s N-terminus might
high-Mr TrxR (Gilberger et al., 1998; Wang et al., 1999). encode a targeting signal that the bacterial expression
This C-terminal redox-active centre of the mammalian system can not deal with. Analysis of the deduced amino
enzyme comprises a selenocysteine-cysteine pair (Se- acid sequence using PlasmoAP (Foth et al., 2003) sug-
CysCys) whereas the Plasmodium TrxR contains a gests that the protein has a signal peptide, but the predic-
CysXXXXCys motif instead (Gilberger et al., 1998; Wang tion programme gave no firm indication of whether it
et al., 1999; Tamura and Stadtman, 2002). This chemical possesses an apicoplast transit peptide. Thus, it will be
difference has been suggested to be sufficient to allow the necessary to verify experimentally the localization of this

thioredoxin-dependent peroxidase, which potentially Mitochondrial redox and antioxidant systems of
might reside within the apicoplast. The gene encoding this P. falciparum
peroxidase appears to be expressed at relatively low
levels during the trophozoite and schizont stages of Energy metabolism in asexual stages of P. falciparum
P. falciparum, suggesting either a secondary role for this apparently does not require an active mitochondrion as
peroxidase for the reduction of hydroperoxides in the par- glucose is mainly metabolized into lactate which is
asites or perhaps that the parasites elevate the expression excreted from the parasite (Jacobasch et al., 1990). Mito-
of the protein if they are under enhanced oxidative or chondrial activities appear to be primarily necessary to
environmental stress (Le Roch et al., 2003). provide a number of essential metabolites such as pyrim-
The cytosolic 2-Cys peroxiredoxin is thioredoxin depen- idines, the biosynthetic pathway of which requires an
dent and efficiently reduces hydrogen peroxide in vitro active electron transport chain (Gutteridge et al., 1979).
with kinetic parameters that suggest a crucial function as The activity of the parasite’s respiratory chain is much
hydroperoxide peroxidase in vivo (Akerman and Müller, lower than in mammalian cells (Fry and Beesley, 1991) but
2003). Its catalytic efficiency for the reduction of H2O2 of it can still be assumed that it leaks superoxide anions that
6.7 ¥ 106 M-1 s-1 qualifies the enzyme as one of the most need to be detoxified. As outlined above, a mitochondrial
efficient peroxiredoxins characterized so far. This perhaps SOD has been identified in P. falciparum but it is unclear
reflects the particular importance of the enzyme for P. how the resulting hydrogen peroxide is reduced within the
falciparum as the parasites lack back-up systems such as organelle. A peroxiredoxin potentially located in the mito-
a genuine glutathione peroxidase (Sztajer et al., 2001) chondrion has recently been described (Rahlfs and
and catalase and so are very probably heavily dependent Becker, 2001) and a potential mitochondrial thioredoxin
on peroxiredoxins for the reduction of ROS. The high has been annotated in the GenBank database (Accession
transcription level of PfTrx-Px1 throughout the erythrocytic No. AAQ05974). However, there is no experimental evi-
development of the parasites is in agreement with this dence that verifies the predicted functions and localization
hypothesis. The gene is also transcribed in the gameto- of both proteins. Even if they are present in the parasite’s
cyte and sporozoite stages of P. falciparum although the mitochondrion, the organelle apparently does not contain
expression level are lower than in the erythrocytic stages TrxR, which initiates the thioredoxin redox cascade.
of the parasites (Le Roch et al., 2003). In spite of this We therefore have postulated that the low-molecular-
potential importance of PfTrx-Px1, the knockout of the weight thiol lipoic acid, an essential cofactor of mitochon-
gene resulted in viable parasites that were only slightly drial a-ketoacid dehydrogenase complexes, provides
more susceptible to oxidative and nitrosative stress than reducing equivalents for the thioredoxin-dependent detox-
the wild-type parasites (Komaki-Yasuda et al., 2003). ification of hydroperoxides (Fig. 4). It has been previously
These results suggest that there must be other efficient shown that the multienzyme complexes support thiore-
hydroperoxide-reducing enzymes present in the parasites doxin reduction (Bunik and Follmann, 1993) and in
that compensate for the loss of PfTrx-Px1 activity – pos- Mycobacterium (Bryk et al. 2002) this mechanism of
sibly the glutathione peroxidase-like protein discussed hydroperoxide detoxification was suggested to be of piv-
above or the 1-Cys peroxiredoxin or GST. Unfortunately, otal importance for the antioxidant defence of the bacteria.
these possibilities were not analysed in the PfTrx-Px1 null Furthermore, the presence of three distinct a-ketoacid
mutants so as yet it is not clear how these mutant para- dehydrogenase complexes has been verified in P. falci-
sites cope with their pro-oxidant environment. parum erythrocytic stages and they were experimentally
The importance of the 1-Cys peroxiredoxin is also shown to be located in the mitochondrion and the apico-
uncertain. The high expression level of the gene encoding plast of the parasites (B. J. Foth et al., unpubl. results; P.
this protein throughout the erythrocytic development (Le J. McMillan et al., unpubl. results). In agreement with this
Roch et al., 2003) suggests that the protein is highly abun- distribution, Plasmodium possesses organelle-specific
dant throughout these parasite stages but its role as a part pathways that supply the parasites with lipoic acid. Lipoic
of the parasite’s antioxidant defence still has to be estab- acid synthesis consisting of lipoic acid synthase and lipoic
lished. The endogenous reducing partner of this protein acid binding protein, responsible for the ligation of newly
has not yet been identified, although there has been a synthesized lipoic acid to the multienzyme complex, are
report that it is reduced by very high concentrations of located in the apicoplast. Lipoic acid protein ligase, the
glutathione (Kawazu et al., 2000). There are a number of enzyme responsible for the attachment of scavenged
studies suggesting cyclophilin, glutathione and GST as lipoic acid to the mitochondrial a-ketoacid dehydrogenase
electron donors for the homologous proteins in other complexes, is localized in the parasite’s mitochondrion
eukaryotes (Chen et al., 2000; Lee et al., 2001; Manevich (Thomsen-Zieger et al., 2003; Wrenger and Müller, 2004)
et al., 2004) and these possibilities still need to be inves- (Fig. 4). Whether mitochondrial lipoic acid is scavenged
tigated for the Plasmodium protein. from the host or whether it is provided to the organelle by

1300 S. Müller
Fig. 4. Hypothetical antioxidant systems in the mitochondrion of Plasmodium falciparum. Plasmodium falciparum erythrocytic stages posses an
active respiratory chain and the enzyme complexes I and III (C1 and C3) contain ubiquinone (UbQ) as cofactor. It is well established that both
complexes leak superoxide anions that have to be detoxified by a mitochondrial SOD (SOD 2). During this reaction H2O2 is generated and this
needs to be reduced to prevent oxidative damage to the organelle. The Plasmodium genome contains genes encoding a second thioredoxin-like
protein (Trx) and a potential mitochondrial peroxiredoxin (Trx-Px) which could be responsible for reducing the hydrogen peroxide formed by the
electron transport chain. However, as the parasites appear to lack a mitochondrial thioredoxin reductase, we postulate an alternative mechanism
by which the thioredoxin redox cycle is driven. Recently we have shown that the parasites possess a lipoic acid protein ligase (LplA) present in
the mitochondrion (Wrenger and Müller, 2004) and it is likely that the metabolite is required to ensure the functionality of the mitochondrial a-
ketoacid dehydrogenase complexes (KADH) by transferring free lipoic acid to their dihydrolipoamide transacylase subunit (E2-subunit). The free
or protein-bound lipoamide provided either by the host or by the lipoic acid synthesis pathway present in the parasite’s apicoplast is taken up
into the mitochondrion by an as yet unidentified mechanism and potentially acts as a reductant of thioredoxin which in turn will reduce the
mitochondrial peroxiredoxin thus guaranteeing the efficient removal of hydroperoxides (ROOH, including H2O2) formed during the metabolic
reactions in the mitochondrion.
the apicoplast localized lipoic acid synthetic pathway the transcriptional and protein levels when erythrocytic
remains for further investigation. However, the fact that stages of P. falciparum were exposed to oxidative stress
Plasmodium possesses the lipoic acid de novo synthesis (Wrenger and Müller, 2003). The role of this enzyme as
pathway only in their plastid organelle might reflect that an integral part of the tricarboxylic acid cycle remains
the metabolite cannot be transported via the extensive uncertain although 2-oxoglutarate clearly could feed into
endomembrane system to other subcellular localizations. the TCA cycle in addition to being used for other biosyn-
In further support of our hypothesis that lipoic acid is thetic and metabolic processes.
involved in the defence against ROS, we have obtained
preliminary result showing that Plasmodium Trx is reduced
Conclusions
by the dihydrolipoamide dehydrogenase/lipoamide redox
system with a second order rate of 1 ¥ 104 M-1 s-1. Despite their almost totally fermentative lifestyle,
Another protein involved in maintaining the mitochondrial intraerythrocytic P. falciparum are heavily dependent on
redox environment is an NADP+-dependent isocitrate efficient antioxidant systems. The major reason for this
dehydrogenase which was shown to be upregulated on requirement is that the parasites ingest host cell haemo-
globin, a source for superoxide anions and haem (FP IX), References
a highly toxic by-product of haemoglobin digestion. These
Akerman, S.E., and Müller, S. (2003) 2-Cys peroxiredoxin
components are the prerequisites for the generation of PfTrx-Px1 is involved in the antioxidant defence of Plasmo-
more toxic ROS via the Fenton reaction (Fig. 2). Taking dium falciparum. Mol Biochem Parasitol 130: 75–81.
into account what we now know about the antioxidant Arner, E.S., and Holmgren, A. (2000) Physiological functions
ability of the intraerythrocytic stages of P. falciparum leads of thioredoxin and thioredoxin reductase. Eur J Biochem
to novel perceptions on the metabolism of the parasite. It 267: 6102–6109.
Aslan, M., Thornley-Brown, D., and Freeman, B.A. (2000)
is possible that the parasites have evolved an almost
Reactive species in sickle cell disease. Ann NY Acad Sci
fermentative lifestyle because they are simply not able to
899: 375–391.
cope with additional oxidative stresses generated by a Aslund, F., and Beckwith, J. (1999) The thioredoxin super-
fully aerobic metabolism (Oliveira and Oliveira, 2002). The family: redundancy, specificity, and gray-area genomics. J
fact that they lack catalase and glutathione peroxidases, Bacteriol 181: 1375–1379.
some of the most important antioxidant enzymes in most Atamna, H., and Ginsburg, H. (1993) Origin of reactive oxy-
eukaryotes, and that infection of the erythrocytes leads to gen species in erythrocytes infected with Plasmodium fal-
ciparum. Mol Biochem Parasitol 61: 231–241.
a reduction of the overall glutathione levels in the infected
Atamna, H., and Ginsburg, H. (1995) Heme degradation in
cell might have been the driving force causing the parasite
the presence of glutathione. A proposed mechanism to
to adopt to this metabolic situation. Notably, the apicom- account for the high levels of non-heme iron found in the
plexan parasite T. gondii contains a cytosolic catalase but membranes of hemoglobinopathic red blood cells. J Biol
the enzyme appears not to be essential for their survival Chem 270: 24876–24883.
(Kwok et al., 2004), suggesting that these related para- Atamna, H., and Ginsburg, H. (1997) The malaria parasite
sites possess efficient back-up systems that allow them supplies glutathione to its host cell – investigation of glu-
tathione transport and metabolism in human erythrocytes
to maintain their intracellular redox balance similar to
infected with Plasmodium falciparum. Eur J Biochem 250:
Plasmodium without catalase. As Plasmodium lacks glu-
670–679.
tathione-dependent peroxidases, it is likely that they rely Atamna, H., Krugliak, M., Shalmiev, G., Deharo, E., Pescar-
solely on the peroxiredoxin-linked detoxification of hydro- mona, G., and Ginsburg, H. (1996) Mode of antimalarial
peroxides whereas glutathione might act as the major low- effect of methylene blue and some of its analogues on
molecular-weight thiol redox buffer and as a detoxification Plasmodium falciparum in culture and their inhibition of P.
metabolite either directly (in the degradation of free FP IX) vinckei petteri and P. yoelii nigeriensis in vivo. Biochem
Pharmacol 51: 693–700.
or as cofactor for GST and glyoxalase. This situation
Ayi, K., Cappadoro, M., Branca, M., Turrini, F., and Arese, P.
clearly renders the parasites highly vulnerable to distur-
(1998) Plasmodium falciparum glutathione metabolism and
bances of their antioxidant systems during their erythro- growth are independent of glutathione system of host
cytic life stages and indeed pro-oxidant drugs such as erythrocyte. FEBS Lett 424: 257–261.
artemisinins that elevate endogenous oxidative stress are Becker, K., Gromer, S., Schirmer, R.H., and Müller, S. (2000)
highly efficient in killing the parasites (Dong and Venner- Thioredoxin reductase as a pathophysiological factor and
strom, 2003). These compounds react with FP IX-Fe2+ drug target. Eur J Biochem 267: 6118–6125.
Becker, K., Rahlfs, S., Nickel, C., and Schirmer, R.H.
present in the parasite cell and are activated by a radical
(2003a) Glutathione – functions and metabolism in the
mechanism. The radical intermediates react with cellular
malarial parasite Plasmodium falciparum. Biol Chem 384:
components and apparently one of the major downstream 551–566.
targets of the drugs is a Ca2+-ATPase (Olliaro et al., 2001; Becker, K., Kanzok, S.M., Iozef, R., Fischer, M., Schirmer,
Meshnick, 2002; Eckstein-Ludwig et al., 2003). The anti- R.H., and Rahlfs, S. (2003b) Plasmoredoxin, a novel
malarial activity of artemisinin can be considered as redox-active protein unique for malarial parasites. Eur J
‘proof-of-concept’ that interfering with the oxidant homeo- Biochem 270: 1057–1064.
Becker, K., Tilley, L., Vennerstrom, J.L., Roberts, D., Roger-
stasis in the parasite will have severe repercussions for
son, S., and Ginsburg, H. (2004) Oxidative stress in
parasite survival. Therefore, it is postulated that the dis-
malaria parasite-infected erythrocytes: host–parasite inter-
ruption of the antioxidant and redox systems of Plasmo- actions. Int J Parasitol 34: 163–189.
dium may be a way to inhibit parasite development during Bender, A., van Dooren, G.G., Ralph, S.A., McFadden, G.I.,
the erythrocytic cycle and thus might offer an excellent and Schneider, G. (2003) Properties and prediction of
way to find new drugs against malaria. mitochondrial transit peptides from Plasmodium falci-
parum. Mol Biochem Parasitol 132: 59–66.
Beppu, M., Ando, K., and Kikugawa, K. (1996) Poly-N-
Acknowledgements acetyllactosaminyl saccharide chains of band 3 as
determinants for anti-band 3 autoantibody binding to
The author would like to thank Professor G.H. Coombs for stim- senescent and oxidized erythrocytes. Cell Mol Biol 42:
ulating discussions and critical reading of the manuscript. S.M. 1007–1024.
is a Wellcome Senior Fellow in Basic Biomedical Science. Birago, C., Pace, T., Picci, L., Pizzi, E., Scotti, R., and Ponzi,

1302 S. Müller
M. (1999) The putative gene for the first enzyme of glu- White, K.P., Barillas-Mury, C., and Kafatos, F.C. (2002)
tathione biosynthesis in Plasmodium berghei and Plasmo- Genome expression analysis of Anopheles gambiae:
dium falciparum. Mol Biochem Parasitol 99: 33–40. responses to injury, bacterial challenge, and malaria infec-
Brydges, S.D., and Carruthers, V.B. (2003) Mutation of an tion. Proc Natl Acad Sci USA 99: 8814–8819.
unusual mitochondrial targeting sequence of SODB2 pro- Dive, D., Gratepanche, S., Year, H., Becuwe, P., Daher, W.,
duces multiple targeting fates in Toxoplasma gondii. J Cell Delplace, P., et al. (2003) Superoxide dismutase in Plas-
Sci 116: 4675–4685. modium: a current survey. Redox Rep 8: 265–267.
Bryk, R., Lima, C.D., Erdjument-Bromage, H., Tempst, P., Dong, Y., and Vennerstrom, J.L. (2003) Mechanisms of in
and Nathan, C. (2002) Metabolic enzymes of mycobacteria situ activation for peroxidic antimalarials. Redox Rep 8:
linked to antioxidant defense by a thioredoxin-like protein. 284–288.
Science 295: 1073–1077. Dubois, V.L., Platel, D.F., Pauly, G., and Tribouley-Duret, J.
Bunik, V.I. (2003) 2-Oxo acid dehydrogenase complexes in (1995) Plasmodium berghei: implication of intracellular glu-
redox regulation. Eur J Biochem 270: 1036–1042. tathione and its related enzyme in chloroquine resistance
Bunik, V., and Follmann, H. (1993) Thioredoxin reduction in vivo. Exp Parasitol 81: 117–124.
dependent on alpha-ketoacid oxidation by alpha-ketoacid Eaton, J.W., Eckman, J.R., Berger, E., and Jacob, H.S.
dehydrogenase complexes. FEBS Lett 336: 197–200. (1976) Suppression of malaria infection by oxidant-
Campanale, N., Nickel, C., Daubenberger, C.A., Wehlan, sensitive host erythrocytes. Nature 264: 758–760.
D.A., Gorman, J.J., Klonis, N., et al. (2003) Identification Eckstein-Ludwig, U., Webb, R.J., Van Goethem, I.D., East,
and characterization of heme-interacting proteins in the J.M., Lee, A.G., Kimura, M., et al. (2003) Artemisinins
malaria parasite, Plasmodium falciparum. J Biol Chem target the SERCA of Plasmodium falciparum. Nature 424:
278: 27354–27361. 957–961.
Cappadoro, M., Giribaldi, G., O’Brien, E., Turrini, F., Mannu, Egan, T.J., Combrinck, J.M., Egan, J., Hearne, G.R.,
F., Ulliers, D., et al. (1998) Early phagocytosis of glucose- Marques, H.M., Ntenteni, S., et al. (2002) Fate of haem
6-phosphate dehydrogenase (G6PD)-deficient erythro- iron in the malaria parasite Plasmodium falciparum. Bio-
cytes parasitized by Plasmodium falciparum may explain chem J 365: 343–347.
malaria protection in G6PD deficiency. Blood 92: 2527– Emanuelsson, O., Henrik Nielsen, H., Brunak, S., and von
2534. Heijne, G. (2000) Predicting subcellular localization of pro-
Chen, J.W., Dodia, C., Feinstein, S.I., Jain, M.K., and Fisher, teins based on their N-terminal amino acid sequence. J
A.B. (2000) 1-Cys peroxiredoxin, a bifunctional enzyme Mol Biol 300: 1005–1016.
with glutathione peroxidase and phospholipase A2 activi- Fairfield, A.S., Meshnick, S.R., and Eaton, J.W. (1983)
ties. J Biol Chem 275: 28421–28427. Malaria parasites adopt host cell superoxide dismutase.
Choi, C.Y., Cerda, J.F., Chu, H.A., Babcock, G.T., and Mar- Science 221: 764–766.
letta, M.A. (1999) Spectroscopic characterization of the Famin, O., Krugliak, M., and Ginsburg, H. (1999) Kinetics of
heme-binding sites in Plasmodium falciparum histidine-rich inhibition of glutathione-mediated degradation of ferriproto-
protein 2. Biochemistry 38: 16916–16924. porphyrin IX by antimalarial drugs. Biochem Pharmacol 58:
Coombs, G.H., Westrop, G.D., Suchan, P., Puzova, G., Hirt, 59–68.
R.P., Embley, T.M., et al. (2004) The amitochondriate Färber, P.M., Becker, K., Müller, S., Schirmer, R.H., and
eukaryote Trichomonas vaginalis contains a divergent Franklin, R.M. (1996) Molecular cloning and characteriza-
thioredoxin-linked peroxiredoxin antioxidant system. J Biol tion of a putative glutathione reductase gene, the PfGR2
Chem 279: 5249–5256. gene, from Plasmodium falciparum. Eur J Biochem 239:
Davioud-Charvet, E., McLeish, M.J., Veine, D.M., Giegel, D., 655–661.
Arscott, L.D., Andricopulo, A.D., et al. (2003) Mechanism- Färber, P.M., Arscott, L.D., Williams, C.H., Jr, Becker, K., and
based inactivation of thioredoxin reductase from Plasmo- Schirmer, R.H. (1998) Recombinant Plasmodium falci-
dium falciparum by Mannich bases. Implication for cytotox- parum glutathione reductase is inhibited by the antimalarial
icity. Biochemistry 42: 13319–13330. dye methylene blue. FEBS Lett 422: 311–314.
Deharo, E., Barkan, D., Krugliak, M., Golenser, J., and Gins- Fernandes, A.P., and Holmgren, A. (2004) Glutaredoxins:
burg, H. (2003) Potentiation of the antimalarial action of glutathione-dependent redox enzymes with functions far
chloroquine in rodent malaria by drugs known to reduce beyond a simple thioredoxin backup system. Antioxid
cellular glutathione levels. Biochem Pharmacol 66: 809– Redox Signal 6: 63–74.
817. Fidock, D.A., Nomura, T., Talley, A.K., Cooper, R.A.,
Denton, H., McGregor, J.C., and Coombs, G.H. (2004) Dzekunov, S.M., Ferdig, M.T., et al. (2000) Mutations in
Reduction of antileishmanial pentavalent antimonial drugs the P. falciparum digestive vacuole transmembrane protein
by a parasite-specific thiol dependent reductase TDR1. PfCRT and evidence for their role in chloroquine resis-
Biochem J 381: 405–412. tance. Mol Cell 6: 861–871.
Destro Bisol, G. (1999) Genetic resistance to malaria, oxida- Filomeni, G., Rotilio, G., and Ciriolo, M.R. (2002) Cell signal-
tive stress and hemoglobin oxidation. Parassitologia 41: ling and the glutathione redox system. Biochem Pharmacol
203–204. 64: 1057–1064.
Di Mascio, P., Murphy, M.E., and Sies, H. (1991) Antioxidant Foth, B.J., Ralph, S.A., Tonkin, C.J., Struck, N.S., Fraunholz,
defense systems: the role of carotenoids, tocopherols, and M., Roos, D.S., et al. (2003) Dissecting apicoplast target-
thiols. Am J Clin Nutr 53: 194S–200S. ing in the malaria parasite Plasmodium falciparum. Sci-
Dimopoulos, G., Christophides, G.K., Meister, S., Schultz, J., ence 299: 705–708.

Frey, P.A. (1997) Radicals in enzymatic reactions. Curr Opin C. (2000) Molecular interactions between Anopheles
Chem Biol 1: 347–356. stephensi midgut cells and Plasmodium berghei: the time
Fridovich, I. (1995) Superoxide radical and superoxide dis- bomb theory of ookinete invasion of mosquitoes. EMBO J
mutases. Annu Rev Biochem 64: 97–112. 19: 6030–6040.
Friedman, M.J. (1978) Erythrocytic mechanism of sickle cell Harwaldt, P., Rahlfs, S., and Becker, K. (2002) Glutathione
resistance to malaria. Proc Natl Acad Sci USA 75: 1994– S-transferase of the malarial parasite Plasmodium falci-
1997. parum: characterization of a potential drug target. Biol
Fritz-Wolf, K., Becker, A., Rahlfs, S., Harwaldt, P., Schirmer, Chem 383: 821–830.
R.H., Kabsch, W., and Becker, K. (2003) X-ray structure of Hirt, R.P., Müller, S., Embley, T.M., and Coombs, G.H.
glutathione S-transferase from the malarial parasite Plas- (2002) The diversity and evolution of thioredoxin reduc-
modium falciparum. Proc Natl Acad Sci USA 100: 13821– tase: new perspectives. Trends Parasitol 18: 302–308.
13826. Hofmann, B., Hecht, H.J., and Flohe, L. (2002) Peroxiredox-
Fry, M., and Beesley, J.E. (1991) Mitochondria of mammalian ins. Biol Chem 383: 347–364.
Plasmodium spp. Parasitology 102: 17–26. Holmgren, A. (2000) Antioxidant function of thioredoxin and
Garrido, E.O., and Grant, C.M. (2002) Role of thioredoxins glutaredoxin systems. Antioxid Redox Signal 2: 811–820.
in the response of Saccharomyces cerevisiae to oxidative Homolya, L., Varadi, A., and Sarkadi, B. (2003) Multidrug
stress induced by hydroperoxides. Mol Microbiol 43: 993– resistance-associated proteins: export pumps for conju-
1003. gates with glutathione, glucuronate or sulfate. Biofactors
Gilberger, T.W., Walter, R.D., and Müller, S. (1997) Identi- 17: 103–114.
fication and characterization of the functional amino Hunt, N.H., and Stocker, R. (1990) Oxidative stress and the
acids at the active site of the large thioredoxin reductase redox status of malaria-infected erythrocytes. Blood Cells
from Plasmodium falciparum. J Biol Chem 272: 29584– 16: 499–526.
29589. Imlay, J.A. (2003) Pathways of oxidative damage. Annu Rev
Gilberger, T.W., Bergmann, B., Walter, R.D., and Müller, S. Microbiol 57: 395–418.
(1998) The role of the C-terminus for catalysis of the large Inoue, M., Sato, E.F., Nishikawa, M., Park, A.M., Kira, Y.,
thioredoxin reductase from Plasmodium falciparum. FEBS Imada, I., and Utsumi, K. (2003) Mitochondrial generation
Lett 425: 407–410. of reactive oxygen species and its role in aerobic life. Curr
Gilberger, T.W., Schirmer, R.H., Walter, R.D., and Müller, S. Med Chem 10: 2495–2505.
(2000) Deletion of the parasite-specific insertions and Jacobasch, G., Buckwitz, D., Gerth, C., and Thamm, R.
mutation of the catalytic triad in glutathione reductase from (1990) Regulation of the energy metabolism of Plasmo-
chloroquine-sensitive Plasmodium falciparum 3D7. Mol dium berghei. Biomed Biochim Acta 49: S289–S294.
Biochem Parasitol 107: 169–179. de Jong, K., Geldwerth and Kuypers, F.A. (1997) Oxidative
Ginsburg, H., Famin, O., Zhang, J., and Krugliak, M. (1998) damage does not alter membrane phospholipid asymmetry
Inhibition of glutathione-dependent degradation of heme by in human erythrocytes. Biochemistry 36: 6768–6776.
chloroquine and amodiaquine as a possible basis for their Kanzok, S.M., Schirmer, R.H., Turbachova, I., Iozef, R., and
antimalarial mode of action. Biochem Pharmacol 56: Becker, K. (2000) The thioredoxin system of the malaria
1305–1313. parasite Plasmodium falciparum. Glutathione reduction
Giribaldi, G., Ulliers, D., Mannu, F., Arese, P., and Turrini, F. revisited. J Biol Chem 275: 40180–40186.
(2001) Growth of Plasmodium falciparum induces stage- Karplus, P.A., and Schulz, G.E. (1987) Refined structure of
dependent haemichrome formation, oxidative aggregation glutathione reductase at 1.54 A resolution. J Mol Biol 195:
of band 3, membrane deposition of complement and anti- 701–729.
bodies, and phagocytosis of parasitized erythrocytes. Br J Kawazu, S., Tsuji, N., Hatabu, T., Kawai, S., Matsumoto, Y.,
Haematol 113: 492–499. and Kano, S. (2000) Molecular cloning and characteriza-
Grant, C.M. (2001) Role of the glutathione/glutaredoxin and tion of a peroxiredoxin from the human malaria parasite
thioredoxin systems in yeast growth and response to stress Plasmodium falciparum. Mol Biochem Parasitol 109: 165–
conditions. Mol Microbiol 39: 533–541. 169.
Gratepanche, S., Menage, S., Touati, D., Wintjens, R., Del- Kita, K., Hirawake, H., Miyadera, H., Amino, H., and Takeo,
place, P., Fontecave, M., et al. (2002) Biochemical and S. (2002) Role of complex II in anaerobic respiration of the
electron paramagnetic resonance study of the iron super- parasite mitochondria from Ascaris suum and Plasmodium
oxide dismutase from Plasmodium falciparum. Mol Bio- falciparum. Biochim Biophys Acta 1553: 123–139.
chem Parasitol 120: 237–246. Komaki-Yasuda, K., Kawazu, S., and Kano, S. (2003) Disrup-
Griffith, O.W. (1999) Biologic and pharmacologic regulation tion of the Plasmodium falciparum 2-Cys peroxiredoxin
of mammalian glutathione synthesis. Free Radic Biol Med gene renders parasites hypersensitive to reactive oxygen
27: 922–935. and nitrogen species. FEBS Lett 547: 140–144.
Gutteridge, W.E., Dave, D., and Richards, W.H. (1979) Con- Krauth-Siegel, R.L., Jockers-Scherubl, M.C., Becker, K., and
version of dihydroorotate to orotate in parasitic protozoa. Schirmer, R.H. (1989) NADPH-dependent disulphide
Biochim Biophys Acta 582: 390–401. reductases. Biochem Soc Trans 17: 315–317.
Guttmann, P., and Ehrlich, P. (1891) Über die Wirkung des Krnajski, Z., Gilberger, T.W., Walter, R.D., and Müller, S.
Methylenblau bei Malaria. Berl Klin Wochenzeitschr 28: (2000) Intersubunit interactions in Plasmodium falciparum
953–956. thioredoxin reductase. J Biol Chem 275: 40874–40878.
Han, Y.S., Thompson, J., Kafatos, F.C., and Barillas-Mury, Krnajski, Z., Gilberger, T.W., Walter, R.D., Cowman, A.F.,

1304 S. Müller
and Müller, S. (2002) Thioredoxin reductase is essential Meierjohann, S., Walter, R.D., and Müller, S. (2002b) Glu-
for the survival of Plasmodium falciparum erythrocytic tathione synthetase from Plasmodium falciparum. Biochem
stages. J Biol Chem 277: 25970–25975. J 363: 833–838.
Krnajski, Z., Gilberger, T.W., Walter, R.D., and Müller, S. Meshnick, S.R. (2002) Artemisinin: mechanisms of action,
(2001a) The malaria parasite Plasmodium falciparum pos- resistance and toxicity. Int J Parasitol 32: 1655–1660.
sesses a functional thioredoxin system. Mol Biochem Par- Motohashi, K., Koyama, F., Nakanishi, Y., Ueoka-Nakanishi,
asitol 112: 219–228. H., and Hisabori, T. (2003) Chloroplast cyclophilin is a
Krnajski, Z., Walter, R.D., and Müller, S. (2001b) Isolation target protein of thioredoxin. Thiol modulation of the pepti-
and functional analysis of two thioredoxin peroxidases dyl-prolyl cis-trans isomerase activity. J Biol Chem 278:
(peroxiredoxins) from Plasmodium falciparum. Mol Bio- 31848–31852.
chem Parasitol 113: 303–308. Mu, J., Ferdig, M.T., Feng, X., Joy, D.A., Duan, J., Furuya,
Kumar, S., Christophides, G.K., Cantera, R., Charles, B., T., et al. (2003) Multiple transporters associated with
Han, Y.S., Meister, S., et al. (2003) The role of reactive malaria parasite responses to chloroquine and quinine.
oxygen species on Plasmodium melanotic encapsulation Mol Microbiol 49: 977–989.
in Anopheles gambiae. Proc Natl Acad Sci USA 100: Müller, S., Liebau, E., Walter, R.D., and Krauth-Siegel, R.L.
14139–14144. (2003) Thiol-based redox metabolism of protozoan para-
Kwok, L.Y., Schlüter, D., Clayton, C., and Soldati, D. (2004) sites. Trends Parasitol 19: 320–328.
The antioxidant systems in Toxoplasma gondii and the role Oliveira, P.L., and Oliveira, M.F. (2002) Vampires, pasteur
of cytosolic catalase in defence against oxidative injury. and reactive oxygen species. Is the switch from aerobic to
Mol Microbiol 51: 47–61. anaerobic metabolism a preventive antioxidant defence in
Le Roch, K.G., Zhou, Y., Blair, P.L., Grainger, M., Moch, J.K., blood-feeding parasites? FEBS Lett 525: 3–6.
Haynes, J.D., et al. (2003) Discovery of gene function by Olliaro, P.L., Haynes, R.K., Meunier, B., and Yuthavong, Y.
expression profiling of the malaria parasite life cycle. Sci- (2001) Possible modes of action of the artemisinin-type
ence 301: 1503–1508. compounds. Trends Parasitol 17: 122–126.
Lee, S.P., Hwang, Y.S., Kim, Y.J., Kwon, K.S., Kim, H.J., Omodeo-Sale, F., Motti, A., Basilico, N., Parapini, S., Olliaro,
Kim, K., and Chae, H.Z. (2001) Cyclophilin a binds to P., and Taramelli, D. (2003) Accelerated senescence of
peroxiredoxins and activates its peroxidase activity. J Biol human erythrocytes cultured with Plasmodium falciparum.
Chem 276: 29826–29832. Blood 102: 705–711.
Liebau, E., Bergmann, B., Campbell, A.M., Teesdale-Spittle, Ouaissi, M.A., Dubremetz, J.F., Schöneck, R., Fernandez-
P., Brophy, P.M., Lüersen, K., and Walter, R.D. (2002) The Gomez, R., Gomez-Corvera, R., Billaut-Mulot, O., et al.
glutathione S-transferase from Plasmodium falciparum. (1995) Trypanosoma cruzi: a 52-kDa protein sharing
Mol Biochem Parasitol 124: 85–90. sequence homology with glutathione S-transferase is local-
Liochev, S.I., and Fridovich, I. (1999) Superoxide and iron: ized in parasite organelles morphologically resembling res-
partners in crime. IUBMB Life 48: 157–161. ervosomes. Exp Parasitol 81: 453–461.
Loria, P., Miller, S., Foley, M., and Tilley, L. (1999) Inhibition Oude Elferink, R.P., Ottenhoff, R., Liefting, W.G., Schoe-
of the peroxidative degradation of haem as the basis of maker, B., Groen, A.K., and Jansen, P.L. (1990) ATP-
action of chloroquine and other quinoline antimalarials. dependent efflux of GSSG and GS-conjugate from isolated
Biochem J 339: 363–370. rat hepatocytes. Am J Physiol 258: G699–G706.
Lüersen, K., Walter, R.D., and Müller, S. (1999) The putative Parker, P.D., Tilley, L., and Klonis, N. (2004) Plasmodium
gamma-glutamylcysteine synthetase from Plasmodium fal- falciparum induces reorganization of host membrane pro-
ciparum contains large insertions and a variable tandem teins during intraerythrocytic growth. Blood 103: 2404–
repeat. Mol Biochem Parasitol 98: 131–142. 2406.
Lüersen, K., Walter, R.D., and Müller, S. (2000) Plasmodium Pasvol, G., Weatherall, D.J., and Wilson, R.J. (1978) Cellular
falciparum-infected red blood cells depend on a functional mechanism for the protective effect of haemoglobin S
glutathione de novo synthesis attributable to an enhanced against P. falciparum malaria. Nature 274: 701–703.
loss of glutathione. Biochem J 346: 545–552. Perbandt, M., Burmeister, C., Walter, R.D., Betzel, C., and
Manevich, Y., Feinstein, S.I., and Fisher, A.B. (2004) Activa- Liebau, E. (2004) Native and inhibited structure of a Mu
tion of the antioxidant enzyme 1-CYS peroxiredoxin class-related glutathione S-transferase from Plasmodium
requires glutathionylation mediated by heterodimerization Falciparum. J Biol Chem 279: 1336–1342.
with pi GST. Proc Natl Acad Sci USA 101: 3780–3785. Perez-Rosado, J., Gervais, G.W., Ferrer-Rodriguez, I.,
Mashima, R., Tilley, L., Siomos, M.A., Papalexis, V., Raftery, Peters, W., and Serrano, A.E. (2002) Plasmodium berghei:
M.J., and Stocker, R. (2002) Plasmodium falciparum histi- analysis of the gamma-glutamylcysteine synthetase gene
dine-rich protein-2 (PfHRP2) modulates the redox activity in drug-resistant lines. Exp Parasitol 101: 175–182.
of ferri-protoporphyrin IX (FePPIX): peroxidase-like activity Platel, D.F., Mangou, F., and Tribouley-Duret, J. (1999) Role
of the PfHRP2-FePPIX complex. J Biol Chem 277: 14514– of glutathione in the detoxification of ferriprotoporphyrin IX
14520. in chloroquine resistant Plasmodium berghei. Mol Biochem
Meierjohann, S., Walter, R.D., and Müller, S. (2002a) Regu- Parasitol 98: 215–223.
lation of intracellular glutathione levels in erythrocytes Rahlfs, S., and Becker, K. (2001) Thioredoxin peroxidases of
infected with chloroquine-sensitive and chloroquine- the malarial parasite Plasmodium falciparum. Eur J
resistant Plasmodium falciparum. Biochem J 368: 761– Biochem 268: 1404–1409.
768. Rahlfs, S., Fischer, M., and Becker, K. (2001) Plasmodium

falciparum possesses a classical glutaredoxin and a sec- Thomsen-Zieger, N., Schachtner, J., and Seeber, F. (2003)
ond, glutaredoxin-like protein with a PICOT homology Apicomplexan parasites contain a single lipoic acid syn-
domain. J Biol Chem 276: 37133–37140. thase located in the plastid. FEBS Lett 547: 80–86.
Rahlfs, S., Nickel, C., Deponte, M., Schirmer, R.H., and Tilley, L., Loria, P., and Floey, M. (2001) Cholorquine and
Becker, K. (2003) Plasmodium falciparum thioredoxins and other quinoline antimalarials. In Antimalarial Chemother-
1
glutaredoxins as central players in redox metabolism. apy. Rosenthal, P.J. (ed.). Totowa, NJ: Humana Press, pp.
Redox Rep 8: 246–250. 87–122.
Reed, M.B., Saliba, K.J., Caruana, S.R., Kirk, K., and Cow- Tran, C.V., and Saier, M.H., Jr (2004) The principal chloro-
man, A.F. (2000) Pgh1 modulates sensitivity and resis- quine resistance protein of Plasmodium falciparum is a
tance to multiple antimalarials in Plasmodium falciparum. member of the drug/metabolite transporter superfamily.
Nature 403: 906–909. Microbiology 150: 1–3.
Ritz, D., and Beckwith, J. (2001) Roles of thiol-redox path- Turrens, J.F. (2004) Oxidative stress and antioxidant
ways in bacteria. Annu Rev Microbiol 55: 21–48. defenses: a target for the treatment of diseases caused by
Roberts, D.J., and Williams, T.N. (2003) Haemoglobi- parasitic protozoa. Mol Aspects Med 25: 211–220.
nopathies and resistance to malaria. Redox Rep 8: 304– Ursos, L.M., and Roepe, P.D. (2002) Chloroquine resistance
310. in the malarial parasite, Plasmodium falciparum. Med Res
Sarma, G.N., Savvides, S.N., Becker, K., Schirmer, M., Rev 22: 465–491.
Schirmer, R.H., and Karplus, P.A. (2003) Glutathione Vickers, T.J., and Fairlamb, A.H. (2004) Trypanothione S-
reductase of the malarial parasite Plasmodium falciparum: transferase activity in a trypanosomatid ribosomal elonga-
crystal structure and inhibitor development. J Mol Biol 328: tion factor 1B. J Biol Chem 279: 27246–27256.
893–907. Wang, P.F., Arscott, L.D., Gilberger, T.W., Müller, S., and
Schirmer, R.H., Müller, J.G., and Krauth-Siegel, R.L. (1995) Williams, C.H., Jr (1999) Thioredoxin reductase from Plas-
Disulfide-reductase inhibitors as chemotherapeutic agents: modium falciparum: evidence for interaction between the
the design of drugs for trypanosomiasis and malaria. C-terminal cysteine residues and the active site disulfide-
Angew Chem Int Ed Engl 34: 141–154. dithiol. Biochemistry 38: 3187–3196.
Shalev, O., and Hebbel, R.P. (1996) Catalysis of soluble Williams, C.H., Jr (1992) Lipoamide dehydrogenase, glu-
hemoglobin oxidation by free iron on sickle red cell mem- tathione reductase, thioredoxin reductase and mercuric ion
branes. Blood 87: 3948–3952. reductase – a family of flavoenzyme transhydrogenases.
Shalev, O., Repka, T., Goldfarb, A., Grinberg, L., Abrahamov, In Chemistry and Biochemistry of Flavoenzymes, Vol. III.
A., Olivieri, N.F., et al. (1995) Deferiprone (L1) chelates Müller, F. (ed.). Boca Raton, FL: CRC Press, pp. 121–211.
pathologic iron deposits from membranes of intact thalas- Williams, C.H., Arscott, L.D., Müller, S., Lennon, B.W., Lud-
saemic and sickle red blood cells both in vitro and in vivo. wig, M.L., Wang, P.F., et al. (2000) Thioredoxin reductase
Blood 86: 2008–2013. two modes of catalysis have evolved. Eur J Biochem 267:
Sienkiewicz, N., Daher, W., Dive, D., Wrenger, C., Viscogli- 6110–6117.
osi, E., Wintjens, R., et al. (2004) Identification of a mito- Williams, T.N., Weatherall, D.J., and Newbold, C.I. (2002)
chondrial superoxide dismutase with an unusual targeting The membrane characteristics of Plasmodium falci-
sequence in Plasmodium falciparum. Mol Biochem Parasi- parum-infected and -uninfected heterozygous alpha (0)
tol (in press). thalassaemic erythrocytes. Br J Haematol 118: 663–
Sies, H. (1993) Strategies of antioxidant defense. Eur J Bio- 670.
chem 215: 213–219. Wong, C.M., Siu, K.L., and Jin, D.Y. (2004) Peroxiredoxin-
Sies, H. (1997) Oxidative stress: oxidants and antioxidants. null yeast cells are hypersensitive to oxidative stress and
Exp Physiol 82: 291–295. genomically unstable. J Biol Chem 279: 23207–23213.
Sies, H. (1999) Glutathione and its role in cellular functions. Wood, Z.A., Schröder, E., Robin Harris, J., and Poole, L.B.
Free Radic Biol Med 27: 916–921. (2003) Structure, mechanism and regulation of peroxire-
Simoes, A.P., van den Berg, J.J., Roelofsen, B., and Op den doxins. Trends Biochem Sci 28: 32–40.
Kamp, J.A. (1992) Lipid peroxidation in Plasmodium Wrenger, C., and Müller, S. (2003) Isocitrate dehydrogenase
falciparum-parasitized human erythrocytes. Arch Biochem of Plasmodium falciparum. Eur J Biochem 270: 1775–
Biophys 298: 651–657. 1783.
Sztajer, H., Gamain, B., Aumann, K.D., Slomianny, C., Wrenger, C., and Müller, S. (2004) The human malaria par-
Becker, K., Brigelius-Flohe, R., and Flohe, L. (2001) The asite Plasmodium falciparum has distinct organelle-
putative glutathione peroxidase gene of Plasmodium falci- specific lipoylation pathways. Mol Microbiol 53: 103–113.
parum codes for a thioredoxin peroxidase. J Biol Chem Yodoi, J., Masutani, H., and Nakamura, H. (2001) Redox
276: 7397–7403. regulation by the human thioredoxin system. Biofactors 15:
Tamura, T., and Stadtman, T.C. (2002) Mammalian thiore- 107–111.
doxin reductases. Methods Enzymol 347: 297–306. Zhang, J., Krugliak, M., and Ginsburg, H. (1999) The fate of
Thomas, J.A., Poland, B., and Honzatko, R. (1995) Protein ferriprotorphyrin IX in malaria infected erythrocytes in con-
sulfhydryls and their role in the antioxidant function of pro- junction with the mode of action of antimalarial drugs. Mol
tein S-thiolation. Arch Biochem Biophys 319: 1–9. Biochem Parasitol 99: 129–141.

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Blackwell Science, LtdOxford, UKMMIMolecular Microbiology0950-382XBlackwell Publishing Ltd, 2004? 200453512911305Review ArticleAntioxidant defence of PlasmodiumS.

Molecular Microbiology (2004) 53(5), 1291–1305 doi:10.1111/j.1365-2958.2004.04257.x

Redox and antioxidant systems of the malaria parasite

Sylke Müller*† superoxide anions, hydrogen peroxide and hydroxyl radi-

© 2004 Blackwell Publishing Ltd

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

Fig. 3. Control of intracellular glutathione

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

Table 1. Genes encoding small redox active proteins in P. falciparum.

Gene Protein Location Predicted localizationa

trx 1 Thioredoxin-WCGPC-motif Chr.14 (PF14_0545) Cytosol

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

© 2004 Blackwell Publishing Ltd, Molecular Microbiology, 53, 1291–1305

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