Plasmodium falciparum--do killers commit suicide?
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Animals↗
Biomedical subjects
Katja Becker
Publications and source records attributed to Katja Becker.
Of malaria, metabolism and membrane transport.
With the sequencing of the Plasmodium falciparum genome now complete, increasing attention is turning to the function of gene products and to cell-regulatory processes. The combination of in silico analyses with modern molecular and biophysical methods is leading to rapid advances in our understanding of the mechanisms underlying the biochemistry and physiology of the parasite and its host cell. In this brief review, we present a "snap shot" of recent work in this area, with particular emphasis on aspects relevant to the development of new antimalarial drugs.
Animals↗
Attention deficits and subclinical epileptiform discharges: are EEG diagnostics in ADHD optional or essential?
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Attention Deficit Disorder with Hyperactivity↗
Internet chat rooms and suicide.
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Adolescent↗
'Attention deficits and subclinical epileptiform discharges: are EEG diagnostics in ADHD optional or essential?'.
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Attention Deficit Disorder with Hyperactivity↗
Glyoxalase I of the malarial parasite Plasmodium falciparum: evidence for subunit fusion.
Recombinant Plasmodium falciparum glyoxalase I (PfGlx I) was characterized as monomeric Zn(2+)-containing enzyme of 44 kDa. The K(M) value of the methylglyoxal-glutathione adduct is 77+/-15 microM, the k(cat) value being 4000 min(-1) at 25 degrees C and pH 7.0. PfGlx I consists of two halves, each of which is homologous to the small 2-domain glyoxalase I of man. Both parts of the pfglx I gene were overexpressed; the C-terminal half of PfGlx I was found to be a stable protein and formed an enzymatically active dimer. These results support the hypothesis of domain-swapping and subunit fusion as mechanisms in glyoxalase I evolution.
Amino Acid Sequence↗
Mechanism-based inactivation of thioredoxin reductase from Plasmodium falciparum by Mannich bases. Implication for cytotoxicity.
Thioredoxin reductase (TrxR) is the homodimeric flavoenzyme that catalyzes reduction of thioredoxin disulfide (Trx). For Plasmodium falciparum, a causative agent of tropical malaria, TrxR is an essential protein which has been validated as a drug target. The high-throughput screening of 350000 compounds has identified Mannich bases as a new class of TrxR mechanism-based inhibitors. During catalysis, TrxR conducts reducing equivalents from the NADPH-reduced flavin to Trx via the two redox-active cysteine pairs, Cys88-Cys93 and Cys535'-Cys540', referred to as N-terminal and C-terminal cysteine pairs. The structures of unsaturated Mannich bases suggested that they could act as bisalkylating agents leading to a macrocycle that involves both C-terminal cysteines of TrxR. To confirm this hypothesis, different Mannich bases possessing one or two electrophilic centers were synthesized and first studied in detail using glutathione as a model thiol. Michael addition of glutathione to the double bond of an unsaturated Mannich base (3a) occurs readily at physiological pH. Elimination of the amino group, promoted by base-catalyzed enolization of the ketone, is followed by addition of a second nucleophile. The intermediate formed in this reaction is an alpha,beta-unsaturated ketone that can react rapidly with a second thiol. When studying TrxR as a target of Mannich bases, we took advantage of the fact that the charge-transfer complex formed between the thiolate of Cys88 and the flavin in the reduced enzyme can be observed spectroscopically. The data show that it is the C-terminal Cys 535'-Cys540' pair rather than the N-terminal Cys88-Cys93 pair that is modified by the inhibitor. Although alkylated TrxR is unable to turn over its natural substrate Trx, it can reduce low M(r) electron acceptors such as methyl methanethiolsulfonate by using its unmodified N-terminal thiols. On the basis of results with chemically distinct Mannich bases, a detailed mechanism for the inactivation of TrxR is proposed.
Alkylating Agents↗
X-ray structure of glutathione S-transferase from the malarial parasite Plasmodium falciparum.
GSTs catalyze the conjugation of glutathione with a wide variety of hydrophobic compounds, generally resulting in nontoxic products that can be readily eliminated. In contrast to many other organisms, the malarial parasite Plasmodium falciparum possesses only one GST isoenzyme (PfGST). This GST is highly abundant in the parasite, its activity was found to be increased in chloroquine-resistant cells, and it has been shown to act as a ligandin for parasitotoxic hemin. Thus, the enzyme represents a promising target for antimalarial drug development. We now have solved the crystal structure of PfGST at a resolution of 1.9 A. The homodimeric protein of 26 kDa per subunit represents a GST form that cannot be assigned to any of the known GST classes. In comparison to other GSTs, and, in particular, to the human isoforms, PfGST possesses a shorter C-terminal section resulting in a more solvent-accessible binding site for the hydrophobic and amphiphilic substrates. The structure furthermore reveals features in this region that could be exploited for the design of specific PfGST inhibitors.
Animals↗
Selenenyl iodide: a new substrate for mammalian thioredoxin reductase.
Areneselenenyl iodide stabilised by internal chelation has been synthesized and evaluated as a substrate of thioredoxin reductase (TrxR). The reactivity of TrxR obtained from human placenta towards selenenyl iodide was found to be much higher than that of the E. coli enzyme, indicating the essential nature of a selenocysteine residue in the active site of the human enzyme. The addition of thioredoxin (Trx) significantly enhanced the TrxR-catalysed reduction of selenenyl iodide 1. These studies on the reduction of a selenenyl iodide by the thioredoxin system suggest that stable selenenyl iodides could be new substrates for human TrxR. The Trx system could act as a cofactor for iodothyronine deiodinase by reducing the selenenyl iodide intermediate in the second-half of the deiodinase catalytic cycle to regenerate the active site. The TrxR-catalysed reduction of 1 was not inhibited by the anti-thyroid drug, PTU, suggesting that the involvement of the Trx system in the deiodinase cycle may be responsible for the insensitivity of certain deiodinases towards clinically useful thiourea drugs.
Escherichia coli↗
Identification and characterization of heme-interacting proteins in the malaria parasite, Plasmodium falciparum.
The degradation of hemoglobin by the malaria parasite, Plasmodium falciparum, produces free ferriprotoporphyrin IX (FP) as a toxic by-product. In the presence of FP-binding drugs such as chloroquine, FP detoxification is inhibited, and the build-up of free FP is thought to be a key mechanism in parasite killing. In an effort to identify parasite proteins that might interact preferentially with FP, we have used a mass spectrometry approach. Proteins that bind to FP immobilized on agarose include P. falciparum glyceraldehyde-3-phosphate dehydrogenase (PfGAPDH), P. falciparum glutathione reductase (PfGR), and P. falciparum protein disulfide isomerase. To examine the potential consequences of FP binding, we have examined the ability of FP to inhibit the activities of GAPDH and GR from P. falciparum and other sources. FP inhibits the enzymic activity of PfGAPDH with a Ki value of 0.2 microm, whereas red blood cell GAPDH is much less sensitive. By contrast, PfGR is more resistant to FP inhibition (Ki > 25 microm) than its human counterpart. We also examined the ability of FP to inhibit the activities of the additional antioxidant enzymes, P. falciparum thioredoxin reductase, which exhibits a Ki value of 1 microm, and P. falciparum glutaredoxin, which shows more moderate sensitivity to FP. The exquisite sensitivity of PfGAPDH to FP may indicate that the glycolytic pathway of the parasite is particularly susceptible to modulation by FP stress. Inhibition of this pathway may drive flux through the pentose phosphate pathway ensuring sufficient production of reducing equivalents to counteract the oxidative stress induced by FP build-up.
Animals↗
Plasmoredoxin, a novel redox-active protein unique for malarial parasites.
Thioredoxins are a group of small redox-active proteins involved in cellular redox regulatory processes as well as antioxidant defense. Thioredoxin, glutaredoxin, and tryparedoxin are members of the thioredoxin superfamily and share structural and functional characteristics. In the malarial parasite, Plasmodium falciparum, a functional thioredoxin and glutathione system have been demonstrated and are considered to be attractive targets for antimalarial drug development. Here we describe the identification and characterization of a novel 22 kDa redox-active protein in P. falciparum. As demonstrated by in silico sequence analyses, the protein, named plasmoredoxin (Plrx), is highly conserved but found exclusively in malarial parasites. It is a member of the thioredoxin superfamily but clusters separately from other members in a phylogenetic tree. We amplified the gene from a gametocyte cDNA library and overexpressed it in E. coli. The purified gene product can be reduced by glutathione but much faster by dithiols like thioredoxin, glutaredoxin, trypanothione and tryparedoxin. Reduced Plrx is active in an insulin-reduction assay and reduces glutathione disulfide with a rate constant of 640 m-1.s-1 at pH 6.9 and 25 degrees C; glutathione-dependent reduction of H2O2 and hydroxyethyl disulfide by Plrx is negligible. Furthermore, plasmoredoxin provides electrons for ribonucleotide reductase, the enzyme catalyzing the first step of DNA synthesis. As demonstrated by Western blotting, the protein is present in blood-stage forms of malarial parasites. Based on these results, plasmoredoxin offers the opportunity to improve diagnostic tools based on PCR or immunological reactions. It may also represent a specific target for antimalarial drug development and is of phylogenetic interest.
Amino Acid Sequence↗
Increased frequency of rolandic spikes in ADHD children.
PURPOSE: Some children with rolandic epilepsy have associated neuropsychiatric deficits resembling symptoms of attention deficit-hyperactivity disorder (ADHD), the most common neurobehavioral disorder of childhood. The clinical overlap between both syndromes has received relatively little attention. The study examines the frequency of rolandic spikes in nonepileptic children with ADHD and compares it with a historic control group of 3,726 normal school-aged children. ADHD patients with and without discharges are compared regarding age at admission, sex, global functioning, and distribution of ADHD subtypes. METHODS: The EEGs of 483 ADHD outpatients between 2 and 16 years meeting diagnostic criteria for ADHD according to Diagnostic and Statistical Manual of Mental Disorders (DSM-IV) were evaluated prospectively. If rolandic spikes were present, separate sleep EEGs were performed to exclude a bioelectrical status epilepticus during slow-wave sleep. RESULTS: Rolandic spikes were detected in the EEGs of 27 children (5.6%; 22 boys and five girls). Seizure rate during follow-up tended to be larger in children with rolandic spikes. No significant differences were found between ADHD patients with and without spikes regarding sex and global functioning. ADHD children with rolandic spikes came to our attention significantly earlier than did children without discharges and tended to exhibit more hyperactive-impulsive symptoms, evidenced in a larger proportion of the diagnosis of ADHD combined type than ADHD inattentive type. CONCLUSIONS: The frequency of rolandic spikes in children with ADHD is significantly higher than expected from epidemiologic studies. The question arises how ADHD symptoms are related to rolandic spikes in this ADHD subgroup. Possibly rolandic discharges or underlying, not fully understood mechanisms of epileptogenesis decrease the vulnerability threshold, advance the onset, or aggravate the course of ADHD.
Action Potentials↗
Plasmodium falciparum thioredoxins and glutaredoxins as central players in redox metabolism.
Over the last few years, an increasing number of different functions have been ascribed to small redox-active proteins like thioredoxins (Trx) and glutaredoxins (Grx). These functions include redox regulation of transcription and translation, antioxidant defence, involvement in protein folding and cellular signalling, and reduction of ribonucleotide reductase. In the malarial parasite Plasmodium falciparum, a classical Trx and a typical Grx have been described as well as a number of Trx- and Grx-like proteins including monothiol glutaredoxins. Furthermore, plasmoredoxin, a redox-active protein related to Trx, has been characterized; plasmoredoxin is unique for malarial parasites, therefore having great potential as diagnostic tool. In this minireview, we summarize the current knowledge on members of the thioredoxin superfamily and their function in the malarial parasite P. falciparum.
Amino Acid Sequence↗
Methylene blue as an antimalarial agent.
Methylene blue has intrinsic antimalarial activity and it can act as a chloroquine sensitizer. In addition, methylene blue must be considered for preventing methemoglobinemia, a serious complication of malarial anemia. As an antiparasitic agent, methylene blue is pleiotropic: it interferes with hemoglobin and heme metabolism in digestive organelles, and it is a selective inhibitor of Plasmodium falciparum glutathione reductase. The latter effect results in glutathione depletion which sensitizes the parasite for chloroquine action. At the Centre de Recherche en Santé de Nouna in Burkina Faso, we study the combination of chloroquine with methylene blue (BlueCQ) as a possible medication for malaria in endemic regions. A pilot study with glucose-6-phosphate dehydrogenase-sufficient adult patients has been conducted recently.
Animals↗
A putative glutathione peroxidase of Drosophila encodes a thioredoxin peroxidase that provides resistance against oxidative stress but fails to complement a lack of catalase activity.
Cellular defense systems against reactive oxygen species (ROS) include thioredoxin reductase (TrxR) and glutathione reductase (GR). They generate sulfhydryl-reducing systems which are coupled to antioxidant enzymes, the thioredoxin and glutathione peroxidases (TPx and GPx). The fruit fly Drosophila lacks a functional GR, suggesting that the thioredoxin system is the major source for recycling glutathione. Whole genome in silico analysis identified two non-selenium containing putative GPx genes. We examined the biochemical characteristics of one of these gene products and found that it lacks GPx activity and functions as a TPx. Transgene-dependent overexpression of the newly identified Glutathione peroxidase homolog with thioredoxin peroxidase activity (Gtpx-1) gene increases resistance to experimentally induced oxidative stress, but does not compensate for the loss of catalase, an enzyme which, like GTPx-1, functions to eliminate hydrogen peroxide. The results suggest that GTPx-1 is part of the Drosophila Trx antioxidant defense system but acts in a genetically distinct pathway or in a different cellular compartment than catalase.
Animals↗
Glutathione--functions and metabolism in the malarial parasite Plasmodium falciparum.
When present as a trophozoite in human erythrocytes, the malarial parasite Plasmodium falciparum exhibits an intense glutathione metabolism. Glutathione plays a role not only in antioxidative defense and in maintaining the reducing environment of the cytosol. Many of the known glutathione-dependent processes are directly related to the specific lifestyle of the parasite. Reduced glutathione (GSH) supports rapid cell growth by providing electrons for deoxyribonucleotide synthesis and it takes part in detoxifying heme, a product of hemoglobin digestion. Free radicals generated in the parasite can be scavenged in reaction sequences involving the thiyl radical GS* as well as the thiolate GS-. As a substrate of glutathione S-transferase, glutathione is conjugated to non-degradable compounds including antimalarial drugs. Furthermore, it is the coenzyme of the glyoxalase system which detoxifies methylglyoxal, a byproduct of the intense glycolysis taking place in the trophozoite. Proteins involved in GSH-dependent processes include glutathione reductase, glutaredoxins, glyoxalase I and II, glutathione S-transferases, and thioredoxins. These proteins, as well as the ATP-dependent enzymes of glutathione synthesis, are studied as factors in the pathophysiology of malaria but also as potential drug targets. Methylene blue, an inhibitor of the structurally known P. falciparum glutathione reductase, appears to be a promising antimalarial medication when given in combination with chloroquine.
Algorithms↗
Mitochondrial and cytoplasmic thioredoxin reductase variants encoded by a single Drosophila gene are both essential for viability.
Defense against oxidative stress in mammals includes the regeneration of the major thiol reductants glutathione and thioredoxin by glutathione reductase and thioredoxin reductase (TrxR), respectively. In contrast, Drosophila, and possibly insects in general, lacks glutathione reductase and must rely solely on the TrxR system. The mammalian TrxRs described so far are selenoproteins that utilize NADPH to reduce protein as well as nonprotein substrates in mitochondria and cytoplasm of cells. We show that a single Drosophila gene, Trxr-1, encodes non-selenocysteine-containing cytoplasmic and mitochondrial TrxR isoforms that differ with respect to their N termini. We generated transcript-specific mutants and used in vivo approaches to explore the biological functions of the two enzyme variants by introducing the corresponding transgenes into different Trxr-1 mutants. The results show that, although the two TrxR isoforms have similar biochemical properties, their biological functions are not interchangeable.
Animals↗
Expression and regulation of thioredoxin reductases and other selenoproteins in bone.
The expression of thioredoxin reductases and other selenoproteins in cells of the bone microenvironment may represent an important means of regulation of bone resorption and remodeling in health and disease. Selenoproteins and their substrates may influence intracellular and extracellular redox-dependent signaling, transcription factor activity, posttranslational modification of proteins, and general or compartmentalized scavenging from ROIs. However, the evaluation of their biological role in bone and their potential in terms of therapeutic approaches is just beginning.
Bone Diseases, Metabolic↗