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Yarlett N, Jarroll EL, Morada M, Lloyd D. Protists: Eukaryotic single-celled organisms and the functioning of their organelles. Adv Microb Physiol 2024; 84:243-307. [PMID: 38821633 DOI: 10.1016/bs.ampbs.2024.02.001] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 06/02/2024]
Abstract
Organelles are membrane bound structures that compartmentalize biochemical and molecular functions. With improved molecular, biochemical and microscopy tools the diversity and function of protistan organelles has increased in recent years, providing a complex panoply of structure/function relationships. This is particularly noticeable with the description of hydrogenosomes, and the diverse array of structures that followed, having hybrid hydrogenosome/mitochondria attributes. These diverse organelles have lost the major, at one time, definitive components of the mitochondrion (tricarboxylic cycle enzymes and cytochromes), however they all contain the machinery for the assembly of Fe-S clusters, which is the single unifying feature they share. The plasticity of organelles, like the mitochondrion, is therefore evident from its ability to lose its identity as an aerobic energy generating powerhouse while retaining key ancestral functions common to both aerobes and anaerobes. It is interesting to note that the apicoplast, a non-photosynthetic plastid that is present in all apicomplexan protozoa, apart from Cryptosporidium and possibly the gregarines, is also the site of Fe-S cluster assembly proteins. It turns out that in Cryptosporidium proteins involved in Fe-S cluster biosynthesis are localized in the mitochondrial remnant organelle termed the mitosome. Hence, different organisms have solved the same problem of packaging a life-requiring set of reactions in different ways, using different ancestral organelles, discarding what is not needed and keeping what is essential. Don't judge an organelle by its cover, more by the things it does, and always be prepared for surprises.
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Affiliation(s)
- Nigel Yarlett
- Haskins Laboratories, Pace University, New York, NY, United States; The Department of Chemistry and Physical Sciences, Pace University, New York, NY, United States.
| | - Edward L Jarroll
- Department of Biological Sciences, CUNY-Lehman College, Bronx, NY, United States
| | - Mary Morada
- Haskins Laboratories, Pace University, New York, NY, United States
| | - David Lloyd
- Schools of Biosciences and Engineering, Cardiff University, Wales, United Kingdom
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2
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Xia C, Xing X, Zhang W, Wang Y, Jin X, Wang Y, Tian M, Ba X, Hao F. Cysteine and homocysteine can be exploited by GPX4 in ferroptosis inhibition independent of GSH synthesis. Redox Biol 2024; 69:102999. [PMID: 38150992 PMCID: PMC10829872 DOI: 10.1016/j.redox.2023.102999] [Citation(s) in RCA: 22] [Impact Index Per Article: 22.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/22/2023] [Revised: 12/14/2023] [Accepted: 12/14/2023] [Indexed: 12/29/2023] Open
Abstract
Ferroptosis is inhibited by glutathione peroxidase 4 (GPX4), an antioxidant enzyme that uses reduced glutathione (GSH) as a cofactor to detoxify lipid hydroperoxides. As a selenoprotein, the core function of GPX4 is the thiol-dependent redox reaction. In addition to GSH, other small molecules such as cysteine and homocysteine also contain thiols; yet, whether GPX4 can exploit cysteine and homocysteine to directly detoxify lipid hydroperoxides and inhibit ferroptosis has not been addressed. In this study, we found that cysteine and homocysteine inhibit ferroptosis in a GPX4-dependent manner. However, cysteine inhibits ferroptosis independent of GSH synthesis, and homocysteine inhibits ferroptosis through non-cysteine and non-GSH pathway. Furthermore, we used molecular docking and GPX4 activity analysis to study the binding patterns and affinity between GPX4 and GSH, cysteine, and homocysteine. We found that besides GSH, cysteine and homocysteine are also able to serve as substrates for GPX4 though the affinities of GPX4 with cysteine and homocysteine are lower than that with GSH. Importantly, GPX family and the GSH synthetase pathway might be asynchronously evolved. When GSH synthetase is absent, for example in Flexibacter, the fGPX exhibits higher affinity with cysteine and homocysteine than GSH. Taken together, the present study provided the understanding of the role of thiol-dependent redox systems in protecting cells from ferroptosis and propose that GSH might be a substitute for cysteine or homocysteine to be used as a cofactor for GPX4 during the evolution of aerobic metabolism.
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Affiliation(s)
- Chaoyi Xia
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Xiyue Xing
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Wenxia Zhang
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Yang Wang
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Xin Jin
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Yang Wang
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China
| | - Meihong Tian
- School of Physical Education, Northeast Normal University, 5268 Renmin Street, Changchun, Jilin, 130024, China.
| | - Xueqing Ba
- Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China.
| | - Fengqi Hao
- School of Physical Education, Northeast Normal University, 5268 Renmin Street, Changchun, Jilin, 130024, China; Key Laboratory of Molecular Epigenetics of Ministry of Education, Northeast Normal University, Changchun, Jilin, 130024, China.
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3
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Tachezy J, Makki A, Hrdý I. The hydrogenosomes of Trichomonas vaginalis. J Eukaryot Microbiol 2022; 69:e12922. [PMID: 35567536 DOI: 10.1111/jeu.12922] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
This review is dedicated to the 50th anniversary of the discovery of hydrogenosomes by Miklós Müller and Donald Lindmark, which we will celebrate the following year. It was a long journey from the first observation of enigmatic rows of granules in trichomonads at the end of the 19th century to their first biochemical characterization in 1973. The key experiments by Müller and Lindmark revealed that the isolated granules contain hydrogen-producing hydrogenase, similar to some anaerobic bacteria-a discovery that gave birth to the field of hydrogenosomes. It is also important to acknowledge the parallel work of the team of Apolena Čerkasovová, Jiří Čerkasov, and Jaroslav Kulda, who demonstrated that these granules, similar to mitochondria, produce ATP. However, the evolutionary origin of hydrogenosomes remained enigmatic until the turn of the millennium, when it was finally accepted that hydrogenosomes and mitochondria evolved from a common ancestor. After a historical introduction, the review provides an overview of hydrogenosome biogenesis, hydrogenosomal protein import, and the relationship between the peculiar structure of membrane translocases and its low inner membrane potential due to the lack of respiratory complexes. Next, it summarizes the current state of knowledge on energy metabolism, the oxygen defense system, and iron/sulfur cluster assembly.
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Affiliation(s)
- Jan Tachezy
- Department of Parasitology, Faculty of Science, Charles University, BIOCEV, Průmyslová 595, 25242 Vestec, Czech Republic
| | - Abhijith Makki
- Department of Parasitology, Faculty of Science, Charles University, BIOCEV, Průmyslová 595, 25242 Vestec, Czech Republic
| | - Ivan Hrdý
- Department of Parasitology, Faculty of Science, Charles University, BIOCEV, Průmyslová 595, 25242 Vestec, Czech Republic
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Oxygen levels are key to understanding "Anaerobic" protozoan pathogens with micro-aerophilic lifestyles. Adv Microb Physiol 2021; 79:163-240. [PMID: 34836611 DOI: 10.1016/bs.ampbs.2021.09.001] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
Publications abound on the physiology, biochemistry and molecular biology of "anaerobic" protozoal parasites as usually grown under "anaerobic" culture conditions. The media routinely used are poised at low redox potentials using techniques that remove O2 to "undetectable" levels in sealed containers. However there is growing understanding that these culture conditions do not faithfully resemble the O2 environments these organisms inhabit. Here we review for protists lacking oxidative energy metabolism, the oxygen cascade from atmospheric to intracellular concentrations and relevant methods of measurements of O2, some well-studied parasitic or symbiotic protozoan lifestyles, their homeodynamic metabolic and redox balances, organism-drug-oxygen interactions, and the present and future prospects for improved drugs and treatment regimes.
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5
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Identification of potential inhibitors of Trichomonas vaginalis iron-containing superoxide dismutase by computer-aided drug design approach. Struct Chem 2021. [DOI: 10.1007/s11224-021-01766-2] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
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Bhakta SB, Moran JA, Mercer F. Neutrophil interactions with the sexually transmitted parasite Trichomonas vaginalis: implications for immunity and pathogenesis. Open Biol 2020; 10:200192. [PMID: 32873151 PMCID: PMC7536067 DOI: 10.1098/rsob.200192] [Citation(s) in RCA: 15] [Impact Index Per Article: 3.0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/30/2020] [Accepted: 08/11/2020] [Indexed: 12/11/2022] Open
Abstract
Trichomoniasis is the third most common sexually transmitted infection in humans and is caused by the protozoan parasite, Trichomonas vaginalis (Tv). Pathogenic outcomes are more common in women and generally include mild vaginitis or cervicitis. However, more serious effects associated with trichomoniasis include adverse reproductive outcomes. Like other infectious agents, pathogenesis from Tv infection is predicted to be the result of both parasite and host factors. At the site of infection, neutrophils are the most abundant immune cells present and probably play key roles in both parasite clearance and inflammatory pathology. Here, we discuss the evidence that neutrophils home to the site of Tv infection, kill the parasite, and that in some circumstances, parasites possibly evade neutrophil-directed killing. In vitro, the parasite is killed by neutrophils using a novel antimicrobial mechanism called trogocytosis, which probably involves both innate and adaptive immunity. While mechanisms of evasion are mostly conjecture at present, the persistence of Tv infections in patients argues strongly for their existence. Additionally, many strains of Tv harbour microbial symbionts Mycoplasma hominis or Trichomonasvirus, which are both predicted to impact neutrophil responses against the parasite. Novel research tools, especially animal models, will help to reveal the true outcomes of many factors involved in neutrophil-Tv interactions during trichomoniasis.
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Affiliation(s)
| | | | - Frances Mercer
- Department of Biological Sciences, California State Polytechnic University, Pomona, CA, USA
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A systematic review of the literature on mechanisms of 5-nitroimidazole resistance in Trichomonas vaginalis. Parasitology 2020; 147:1383-1391. [PMID: 32729451 DOI: 10.1017/s0031182020001237] [Citation(s) in RCA: 18] [Impact Index Per Article: 3.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/13/2022]
Abstract
BACKGROUND Trichomonas vaginalis is the most common non-viral sexually transmitted infection. 5-Nitroimidazoles [metronidazole (MTZ) and tinidazole (TDZ)] are FDA-approved treatments. To better understand treatment failure, we conducted a systematic review on mechanisms of 5-nitroimidazole resistance. METHODS PubMed, ScienceDirect and EMBASE databases were searched using keywords Trichomonas vaginalis, trichomoniasis, 5-nitroimidazole, metronidazole, tinidazole and drug resistance. Non-English language articles and articles on other treatments were excluded. RESULTS The search yielded 606 articles, of which 550 were excluded, leaving 58 articles. Trichomonas vaginalis resistance varies and is higher with MTZ (2.2-9.6%) than TDZ (0-2%). Resistance can be aerobic or anaerobic and is relative rather than absolute. Differential expression of enzymes involved in trichomonad energy production and antioxidant defenses affects 5-nitroimidazole drug activation; reduced expression of pyruvate:ferredoxin oxidoreductase, ferredoxin, nitroreductase, hydrogenase, thioredoxin reductase and flavin reductase are implicated in drug resistance. Trichomonas vaginalis infection with Mycoplasma hominis or T. vaginalis virus has also been associated with resistance. Trichomonas vaginalis has two genotypes, with greater resistance seen in type 2 (vs type 1) populations. DISCUSSION 5-Nitroimidazole resistance results from differential expression of enzymes involved in energy production or antioxidant defenses, along with genetic mutations in the T. vaginalis genome. Alternative treatments outside of the 5-nitroimidazole class are needed.
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Kutsumura N, Koyama Y, Saitoh T, Yamamoto N, Nagumo Y, Miyata Y, Hokari R, Ishiyama A, Iwatsuki M, Otoguro K, Ōmura S, Nagase H. Structure-Activity Relationship between Thiol Group-Trapping Ability of Morphinan Compounds with a Michael Acceptor and Anti-Plasmodium falciparum Activities. Molecules 2020; 25:molecules25051112. [PMID: 32131542 PMCID: PMC7179212 DOI: 10.3390/molecules25051112] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/24/2020] [Revised: 02/28/2020] [Accepted: 02/28/2020] [Indexed: 11/26/2022] Open
Abstract
7-Benzylidenenaltrexone (BNTX) and most of its derivatives showed in vitro antimalarial activities against chloroquine-resistant and -sensitive Plasmodium falciparum strains (K1 and FCR3, respectively). In addition, the time-dependent changes of the addition reactions of the BNTX derivatives with 1-propanethiol were examined by 1H-NMR experiments to estimate their thiol group-trapping ability. The relative chemical reactivity of the BNTX derivatives to trap the thiol group of 1-propanethiol was correlated highly with the antimalarial activity. Therefore, the measurements of the thiol group-trapping ability of the BNTX derivatives with a Michael acceptor is expected to become an alternative method for in vitro malarial activity and related assays.
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Affiliation(s)
- Noriki Kutsumura
- International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan; (N.K.); (T.S.); (N.Y.); (Y.N.)
- Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8571, Japan;
| | - Yasuaki Koyama
- Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8571, Japan;
| | - Tsuyoshi Saitoh
- International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan; (N.K.); (T.S.); (N.Y.); (Y.N.)
| | - Naoshi Yamamoto
- International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan; (N.K.); (T.S.); (N.Y.); (Y.N.)
| | - Yasuyuki Nagumo
- International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan; (N.K.); (T.S.); (N.Y.); (Y.N.)
| | - Yoshiyuki Miyata
- School of Medicine, Keio University, 35, Shinanomachi, Shinjuku, Tokyo 160-8582, Japan;
| | - Rei Hokari
- Kitasato Institute for Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan; (R.H.); (A.I.); (M.I.); (K.O.); (S.Ō.)
| | - Aki Ishiyama
- Kitasato Institute for Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan; (R.H.); (A.I.); (M.I.); (K.O.); (S.Ō.)
| | - Masato Iwatsuki
- Kitasato Institute for Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan; (R.H.); (A.I.); (M.I.); (K.O.); (S.Ō.)
| | - Kazuhiko Otoguro
- Kitasato Institute for Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan; (R.H.); (A.I.); (M.I.); (K.O.); (S.Ō.)
| | - Satoshi Ōmura
- Kitasato Institute for Life Sciences, Kitasato University, 5-9-1 Shirokane, Minato-ku, Tokyo 108-8641, Japan; (R.H.); (A.I.); (M.I.); (K.O.); (S.Ō.)
| | - Hiroshi Nagase
- International Institute for Integrative Sleep Medicine (WPI-IIIS), University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8575, Japan; (N.K.); (T.S.); (N.Y.); (Y.N.)
- Graduate School of Pure and Applied Sciences, University of Tsukuba, 1-1-1 Tennodai, Tsukuba, Ibaraki 305-8571, Japan;
- Correspondence: ; Tel.: +81-29-853-6437
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9
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Verdaguer IB, Zafra CA, Crispim M, Sussmann RA, Kimura EA, Katzin AM. Prenylquinones in Human Parasitic Protozoa: Biosynthesis, Physiological Functions, and Potential as Chemotherapeutic Targets. Molecules 2019; 24:molecules24203721. [PMID: 31623105 PMCID: PMC6832408 DOI: 10.3390/molecules24203721] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/05/2019] [Revised: 09/25/2019] [Accepted: 10/01/2019] [Indexed: 12/19/2022] Open
Abstract
Human parasitic protozoa cause a large number of diseases worldwide and, for some of these diseases, there are no effective treatments to date, and drug resistance has been observed. For these reasons, the discovery of new etiological treatments is necessary. In this sense, parasitic metabolic pathways that are absent in vertebrate hosts would be interesting research candidates for the identification of new drug targets. Most likely due to the protozoa variability, uncertain phylogenetic origin, endosymbiotic events, and evolutionary pressure for adaptation to adverse environments, a surprising variety of prenylquinones can be found within these organisms. These compounds are involved in essential metabolic reactions in organisms, for example, prevention of lipoperoxidation, participation in the mitochondrial respiratory chain or as enzymatic cofactors. This review will describe several prenylquinones that have been previously characterized in human pathogenic protozoa. Among all existing prenylquinones, this review is focused on ubiquinone, menaquinone, tocopherols, chlorobiumquinone, and thermoplasmaquinone. This review will also discuss the biosynthesis of prenylquinones, starting from the isoprenic side chains to the aromatic head group precursors. The isoprenic side chain biosynthesis maybe come from mevalonate or non-mevalonate pathways as well as leucine dependent pathways for isoprenoid biosynthesis. Finally, the isoprenic chains elongation and prenylquinone aromatic precursors origins from amino acid degradation or the shikimate pathway is reviewed. The phylogenetic distribution and what is known about the biological functions of these compounds among species will be described, as will the therapeutic strategies associated with prenylquinone metabolism in protozoan parasites.
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Affiliation(s)
- Ignasi B. Verdaguer
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Camila A. Zafra
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Marcell Crispim
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Rodrigo A.C. Sussmann
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
- Centro de Formação em Ciências Ambientais, Universidade Federal do Sul da Bahia, Porto Seguro 45810-000 Bahia, Brazil
| | - Emília A. Kimura
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
| | - Alejandro M. Katzin
- Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo, São Paulo 05508000, Brazil; (I.B.V.); (C.A.Z.); (M.C.); (E.A.K.)
- Correspondence: ; Tel.: +55-11-3091-7330; Fax: +5511-3091-7417
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Zimorski V, Mentel M, Tielens AGM, Martin WF. Energy metabolism in anaerobic eukaryotes and Earth's late oxygenation. Free Radic Biol Med 2019; 140:279-294. [PMID: 30935869 PMCID: PMC6856725 DOI: 10.1016/j.freeradbiomed.2019.03.030] [Citation(s) in RCA: 26] [Impact Index Per Article: 4.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Figures] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/15/2018] [Revised: 03/21/2019] [Accepted: 03/26/2019] [Indexed: 01/09/2023]
Abstract
Eukaryotes arose about 1.6 billion years ago, at a time when oxygen levels were still very low on Earth, both in the atmosphere and in the ocean. According to newer geochemical data, oxygen rose to approximately its present atmospheric levels very late in evolution, perhaps as late as the origin of land plants (only about 450 million years ago). It is therefore natural that many lineages of eukaryotes harbor, and use, enzymes for oxygen-independent energy metabolism. This paper provides a concise overview of anaerobic energy metabolism in eukaryotes with a focus on anaerobic energy metabolism in mitochondria. We also address the widespread assumption that oxygen improves the overall energetic state of a cell. While it is true that ATP yield from glucose or amino acids is increased in the presence of oxygen, it is also true that the synthesis of biomass costs thirteen times more energy per cell in the presence of oxygen than in anoxic conditions. This is because in the reaction of cellular biomass with O2, the equilibrium lies very far on the side of CO2. The absence of oxygen offers energetic benefits of the same magnitude as the presence of oxygen. Anaerobic and low oxygen environments are ancient. During evolution, some eukaryotes have specialized to life in permanently oxic environments (life on land), other eukaryotes have remained specialized to low oxygen habitats. We suggest that the Km of mitochondrial cytochrome c oxidase of 0.1-10 μM for O2, which corresponds to about 0.04%-4% (avg. 0.4%) of present atmospheric O2 levels, reflects environmental O2 concentrations that existed at the time that the eukaryotes arose.
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Affiliation(s)
- Verena Zimorski
- Institute of Molecular Evolution, Heinrich-Heine-University, 40225, Düsseldorf, Germany.
| | - Marek Mentel
- Department of Biochemistry, Faculty of Natural Sciences, Comenius University in Bratislava, 851 04, Bratislava, Slovakia.
| | - Aloysius G M Tielens
- Department of Medical Microbiology and Infectious Diseases, Erasmus Medical Center Rotterdam, The Netherlands; Department of Biochemistry and Cell Biology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
| | - William F Martin
- Institute of Molecular Evolution, Heinrich-Heine-University, 40225, Düsseldorf, Germany.
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11
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Boro P, Sultana A, Mandal K, Chattopadhyay S. Transcriptomic changes under stress conditions with special reference to glutathione contents. THE NUCLEUS 2018. [DOI: 10.1007/s13237-018-0256-5] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 10/27/2022] Open
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12
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Monoyios A, Hummel K, Nöbauer K, Patzl M, Schlosser S, Hess M, Bilic I. An Alliance of Gel-Based and Gel-Free Proteomic Techniques Displays Substantial Insight Into the Proteome of a Virulent and an Attenuated Histomonas meleagridis Strain. Front Cell Infect Microbiol 2018; 8:407. [PMID: 30505807 PMCID: PMC6250841 DOI: 10.3389/fcimb.2018.00407] [Citation(s) in RCA: 8] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 07/27/2018] [Accepted: 10/30/2018] [Indexed: 12/29/2022] Open
Abstract
The unicellular protozoan Histomonas meleagridis is notorious for being the causative agent of histomonosis, which can cause high mortality in turkeys and substantial production losses in chickens. The complete absence of commercially available curative strategies against the disease renders the devising of novel approaches a necessity. A fundamental step toward this objective is to understand the flagellate's virulence and attenuation mechanisms. For this purpose we have previously conducted a comparative proteomic analysis of an in vitro cultivated virulent and attenuated histomonad parasite using two-dimensional electrophoresis and MALDI-TOF/TOF. The current work aimed to substantially extend the knowledge of the flagellate's proteome by applying 2D-DIGE and sequential window acquisition of all theoretical mass spectra (SWATH) MS as tools on the two well-defined strains. In the gel-based experiments, 49 identified protein spots were found to be differentially expressed, of which 37 belonged to the in vitro cultivated virulent strain and 12 to the attenuated one. The most frequently identified proteins in the virulent strain take part in cytoskeleton formation, carbohydrate metabolism and adaptation to stress. However, post-translationally modified or truncated ubiquitous cellular proteins such as actin and GAPDH were identified as upregulated in multiple gel positions. This indicated their contribution to processes not related to cytoskeleton and carbohydrate metabolism, such as fibronectin or plasminogen binding. Proteins involved in cell division and cytoskeleton organization were frequently observed in the attenuated strain. The findings of the gel-based studies were supplemented by the gel-free SWATH MS analysis, which identified and quantified 42 significantly differentially regulated proteins. In this case proteins with peptidase activity, metabolic proteins and actin-regulating proteins were the most frequent findings in the virulent strain, while proteins involved in hydrogenosomal carbohydrate metabolism dominated the results in the attenuated one.
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Affiliation(s)
- Andreas Monoyios
- Clinic for Poultry and Fish Medicine, Department for Farm Animals and Veterinary Public Health, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Karin Hummel
- VetCore Facility for Research, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Katharina Nöbauer
- VetCore Facility for Research, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Martina Patzl
- Department for Pathobiology, Institute of Immunology, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Sarah Schlosser
- VetCore Facility for Research, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Michael Hess
- Clinic for Poultry and Fish Medicine, Department for Farm Animals and Veterinary Public Health, University of Veterinary Medicine Vienna, Vienna, Austria
- Christian Doppler Laboratory for Innovative Poultry Vaccines, University of Veterinary Medicine Vienna, Vienna, Austria
| | - Ivana Bilic
- Clinic for Poultry and Fish Medicine, Department for Farm Animals and Veterinary Public Health, University of Veterinary Medicine Vienna, Vienna, Austria
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13
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Leitsch D, Williams CF, Hrdý I. Redox Pathways as Drug Targets in Microaerophilic Parasites. Trends Parasitol 2018; 34:576-589. [PMID: 29807758 DOI: 10.1016/j.pt.2018.04.007] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/29/2018] [Revised: 04/27/2018] [Accepted: 04/28/2018] [Indexed: 01/06/2023]
Abstract
The microaerophilic parasites Entamoeba histolytica, Trichomonas vaginalis, and Giardia lamblia jointly cause hundreds of millions of infections in humans every year. Other microaerophilic parasites such as Tritrichomonas foetus and Spironucleus spp. pose a relevant health problem in veterinary medicine. Unfortunately, vaccines against these pathogens are unavailable, but their microaerophilic lifestyle opens opportunities for specifically developed chemotherapeutics. In particular, their high sensitivity towards oxygen can be exploited by targeting redox enzymes. This review focusses on the redox pathways of microaerophilic parasites and on drugs, either already in use or currently in the state of development, which target these pathways.
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Affiliation(s)
- David Leitsch
- Institute for Specific Prophylaxis and Tropical Medicine, Center for Pathophysiology, Infectiology, and Immunology, Medical University of Vienna, Austria.
| | - Catrin F Williams
- School of Engineering, Cardiff University, Cardiff, Wales, United Kingdom
| | - Ivan Hrdý
- Department of Parasitology, Charles University, Faculty of Science, Prague, Czech Republic
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14
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Westrop GD, Wang L, Blackburn GJ, Zhang T, Zheng L, Watson DG, Coombs GH. Metabolomic profiling and stable isotope labelling of Trichomonas vaginalis and Tritrichomonas foetus reveal major differences in amino acid metabolism including the production of 2-hydroxyisocaproic acid, cystathionine and S-methylcysteine. PLoS One 2017; 12:e0189072. [PMID: 29267346 PMCID: PMC5739422 DOI: 10.1371/journal.pone.0189072] [Citation(s) in RCA: 19] [Impact Index Per Article: 2.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/09/2017] [Accepted: 11/17/2017] [Indexed: 11/19/2022] Open
Abstract
Trichomonas vaginalis and Tritrichomonas foetus are pathogens that parasitise, respectively, human and bovine urogenital tracts causing disease. Using LC-MS, reference metabolomic profiles were obtained for both species and stable isotope labelling with D-[U-13C6] glucose was used to analyse central carbon metabolism. This facilitated a comparison of the metabolic pathways of T. vaginalis and T. foetus, extending earlier targeted biochemical studies. 43 metabolites, whose identities were confirmed by comparison of their retention times with authentic standards, occurred at more than 3-fold difference in peak intensity between T. vaginalis and T. foetus. 18 metabolites that were removed from or released into the medium during growth also showed more than 3-fold difference between the species. Major differences were observed in cysteine and methionine metabolism in which homocysteine, produced as a bi-product of trans-methylation, is catabolised by methionine γ-lyase in T. vaginalis but converted to cystathionine in T. foetus. Both species synthesise methylthioadenosine by an unusual mechanism, but it is not used as a substrate for methionine recycling. T. vaginalis also produces and exports high levels of S-methylcysteine, whereas only negligible levels were found in T. foetus which maintains significantly higher intracellular levels of cysteine. 13C-labeling confirmed that both cysteine and S-methylcysteine are synthesised by T. vaginalis; S-methylcysteine can be generated by recombinant T. vaginalis cysteine synthase using phosphoserine and methanethiol. T. foetus contained higher levels of ornithine and citrulline than T. vaginalis and exported increased levels of putrescine, suggesting greater flux through the arginine dihydrolase pathway. T. vaginalis produced and exported hydroxy acid derivatives of certain amino acids, particularly 2-hydroxyisocaproic acid derived from leucine, whereas negligible levels of these metabolites occurred in T. foetus.
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Affiliation(s)
- Gareth D. Westrop
- Strathclyde Institute of Pharmacy and Biomedical Science, Strathclyde University, Glasgow, United Kingdom
- * E-mail:
| | - Lijie Wang
- Strathclyde Institute of Pharmacy and Biomedical Science, Strathclyde University, Glasgow, United Kingdom
| | | | - Tong Zhang
- Institute of Cancer Sciences, University of Glasgow, Glasgow, United Kingdom
| | - Liang Zheng
- Pediatric Translational Medicine Institute, Shanghai Children’s Medical Center, Shanghai, China
| | - David G. Watson
- Strathclyde Institute of Pharmacy and Biomedical Science, Strathclyde University, Glasgow, United Kingdom
| | - Graham H. Coombs
- Strathclyde Institute of Pharmacy and Biomedical Science, Strathclyde University, Glasgow, United Kingdom
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15
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Mazumdar R, Endler L, Monoyios A, Hess M, Bilic I. Establishment of a de novo Reference Transcriptome of Histomonas meleagridis Reveals Basic Insights About Biological Functions and Potential Pathogenic Mechanisms of the Parasite. Protist 2017; 168:663-685. [DOI: 10.1016/j.protis.2017.09.004] [Citation(s) in RCA: 13] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/07/2017] [Revised: 08/21/2017] [Accepted: 09/23/2017] [Indexed: 12/28/2022]
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16
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Wang SE, Brooks AES, Cann B, Simoes-Barbosa A. The fluorescent protein iLOV outperforms eGFP as a reporter gene in the microaerophilic protozoan Trichomonas vaginalis. Mol Biochem Parasitol 2017; 216:1-4. [PMID: 28602728 DOI: 10.1016/j.molbiopara.2017.06.003] [Citation(s) in RCA: 14] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/03/2017] [Revised: 06/06/2017] [Accepted: 06/07/2017] [Indexed: 01/31/2023]
Abstract
Trichomonas vaginalis is a flagellated protozoan causing a notorious urogenital infection in humans. Due to its anaerobic metabolism, an alternative fluorescent protein that can be readily expressed in oxygen-deprived conditions is ideal. This study assessed the performance of iLOV, which does not require oxygen to function, as compared to the conventional enhanced green fluorescent protein (eGFP) in T. vaginalis. The results indicated that iLOV outperforms eGFP in both transient and stable expression, being detectable earlier and producing higher fluorescent intensity than eGFP in T. vaginalis. This finding facilitates forthcoming genetic studies that will advance the knowledge on this human parasitic infection.
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Affiliation(s)
- Shuqi E Wang
- School of Biological Sciences, University of Auckland, 3A Symonds Street, Auckland 1010, New Zealand
| | - Anna E S Brooks
- School of Biological Sciences, University of Auckland, 3A Symonds Street, Auckland 1010, New Zealand; Maurice Wilkins Centre, University of Auckland, 3A Symonds Street, Auckland 1010, New Zealand
| | - Bronwyn Cann
- School of Biological Sciences, University of Auckland, 3A Symonds Street, Auckland 1010, New Zealand
| | - Augusto Simoes-Barbosa
- School of Biological Sciences, University of Auckland, 3A Symonds Street, Auckland 1010, New Zealand.
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17
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Korosh T, Bujans E, Morada M, Karaalioglu C, Vanden Eynde JJ, Mayence A, Huang TL, Yarlett N. Potential of bisbenzimidazole-analogs toward metronidazole-resistant Trichomonas vaginalis
isolates. Chem Biol Drug Des 2017; 90:489-495. [DOI: 10.1111/cbdd.12972] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2017] [Revised: 02/23/2017] [Accepted: 02/24/2017] [Indexed: 11/28/2022]
Affiliation(s)
- Travis Korosh
- Department of Chemistry and Physical Sciences; Pace University; New York NY USA
- Haskins Laboratories; Pace University; New York NY USA
| | - Emmanuel Bujans
- Department of Chemistry and Physical Sciences; Pace University; New York NY USA
- Haskins Laboratories; Pace University; New York NY USA
| | - Mary Morada
- Haskins Laboratories; Pace University; New York NY USA
| | - Canan Karaalioglu
- Department of Chemistry and Physical Sciences; Pace University; New York NY USA
| | - Jean Jacques Vanden Eynde
- Division of Basic Pharmaceutical Sciences; College of Pharmacy; Xavier University of Louisiana; New Orleans LA USA
| | - Annie Mayence
- Division of Basic Pharmaceutical Sciences; College of Pharmacy; Xavier University of Louisiana; New Orleans LA USA
| | - Tien L. Huang
- Division of Basic Pharmaceutical Sciences; College of Pharmacy; Xavier University of Louisiana; New Orleans LA USA
| | - Nigel Yarlett
- Department of Chemistry and Physical Sciences; Pace University; New York NY USA
- Haskins Laboratories; Pace University; New York NY USA
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18
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Nobre LS, Meloni D, Teixeira M, Viscogliosi E, Saraiva LM. Trichomonas vaginalis Repair of Iron Centres Proteins: The Different Role of Two Paralogs. Protist 2016; 167:222-33. [PMID: 27124376 DOI: 10.1016/j.protis.2016.03.001] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 11/19/2015] [Revised: 01/28/2016] [Accepted: 03/15/2016] [Indexed: 01/04/2023]
Abstract
Trichomonas vaginalis, the causative parasite of one of the most prevalent sexually transmitted diseases is, so far, the only protozoan encoding two putative Repair of Iron Centres (RIC) proteins. Homologs of these proteins have been shown to protect bacteria from the chemical stress imposed by mammalian immunity. In this work, the biochemical and functional characterisation of the T. vaginalis RICs revealed that the two proteins have different properties. Expression of ric1 is induced by nitrosative stress but not by hydrogen peroxide, while ric2 transcription remained unaltered under similar conditions. T. vaginalis RIC1 contains a di-iron centre, but RIC2 apparently does not. Only RIC1 resembles bacterial RICs on spectroscopic profiling and repairing ability of oxidatively-damaged iron-sulfur clusters. Unexpectedly, RIC2 was found to bind DNA plasmid and T. vaginalis genomic DNA, a function proposed to be related with its leucine zipper domain. The two proteins also differ in their cellular localization: RIC1 is expressed in the cytoplasm only, and RIC2 occurs both in the nucleus and cytoplasm. Therefore, we concluded that the two RIC paralogs have different roles in T. vaginalis, with RIC2 showing an unprecedented DNA binding ability when compared with all other until now studied RICs.
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Affiliation(s)
- Lígia S Nobre
- Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av. da República (EAN), 2780-157 Oeiras, Portugal
| | - Dionigia Meloni
- University Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019 - UMR 8204 - CIIL - Centre d'Infection et d'Immunité de Lille, 1 rue du Professeur Calmette, BP 245, 59019 Lille Cedex, France
| | - Miguel Teixeira
- Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av. da República (EAN), 2780-157 Oeiras, Portugal
| | - Eric Viscogliosi
- University Lille, CNRS, Inserm, CHU Lille, Institut Pasteur de Lille, U1019 - UMR 8204 - CIIL - Centre d'Infection et d'Immunité de Lille, 1 rue du Professeur Calmette, BP 245, 59019 Lille Cedex, France
| | - Lígia M Saraiva
- Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Av. da República (EAN), 2780-157 Oeiras, Portugal.
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19
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Structural investigation and inhibitory response of halide on phosphoserine aminotransferase from Trichomonas vaginalis. Biochim Biophys Acta Gen Subj 2016; 1860:1508-18. [PMID: 27102280 DOI: 10.1016/j.bbagen.2016.04.013] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/12/2015] [Revised: 04/04/2016] [Accepted: 04/17/2016] [Indexed: 11/20/2022]
Abstract
BACKGROUND Phosphoserine aminotransferase (PSAT) catalyses the second reversible step of the phosphoserine biosynthetic pathway in Trichomonas vaginalis, which is crucial for the synthesis of serine and cysteine. METHODS PSAT from T. vaginalis (TvPSAT) was analysed using X-ray crystallography, enzyme kinetics, and molecular dynamics simulations. RESULTS The crystal structure of TvPSAT was determined to 2.15Å resolution, and is the first protozoan PSAT structure to be reported. The active site of TvPSAT structure was found to be in a closed conformation, and at the active site PLP formed an internal aldimine linkage to Lys 202. In TvPSAT, Val 340 near the active site while it is Arg in most other members of the PSAT family, might be responsible in closing the active site. Kinetic studies yielded Km values of 54 μM and 202 μM for TvPSAT with OPLS and AKG, respectively. Only iodine inhibited the TvPSAT activity while smaller halides could not inhibit. CONCLUSION Results from the structure, comparative molecular dynamics simulations, and the inhibition studies suggest that iodine is the only halide that can bind TvPSAT strongly and may thus inhibit the activity of TvPSAT. The long loop between β8 and α8 at the opening of the TvPSAT active site cleft compared to other PSATs, suggests that this loop may help control the access of substrates to the TvPSAT active site and thus influences the enzyme kinetics. GENERAL SIGNIFICANCE Our structural and functional studies have improved our understanding of how PSAT helps this organism persists in the environment.
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20
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Patel ND, Lawrence R, Peteroy-Kelly MA. Persistent Mycobacterium bovis-BCG is resistant to glutathione induced reductive stress killing. Microb Pathog 2016; 95:124-132. [PMID: 26997649 DOI: 10.1016/j.micpath.2016.03.004] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/08/2016] [Revised: 03/03/2016] [Accepted: 03/07/2016] [Indexed: 10/22/2022]
Abstract
This study focuses on the redox stress response in mycobacteria elicited by a host-derived, thiol-based detoxification molecule, glutathione (GSH). Although the growth and viability of Mycobacterium bovis-BCG (BCG) was hampered by exposure to 8 mM GSH, oxygen depleted, persistent BCG (NRP BCG) resisted GSH-mediated killing. Fast growing mycobacteria also resisted GSH-mediated killing. To determine the mechanisms behind these observations, we evaluated the levels of intracellular ATP in both BCG and NRP BCG exposed to 8 mM GSH. Intracellular ATP levels increased from 0.13 to 2.3 μM in BCG upon exposure to GSH. The levels of ATP remained low and unchanged when NRP BCG was exposed to GSH. Using both HPLC and a cell-based thiol detection assay, it was determined that GSH stimulates the production of mycothiol (MSH) by BCG approximately 5.7 fold. The levels of MSH did not change upon exposure of NRP BCG to GSH. MSH is an alternative, thiol-based detoxification molecule employed by mycobacteria. Changes in the cytoplasmic concentrations of this molecule are suggestive of redox imbalances. Together, GSH and MSH may introduce excess reducing equivalents into the mycobacterial cytoplasm; leading to reductive stress. The modulation of NAD(+) levels through alterations in ATP metabolism can enhance the cells ability to bind excess reducing equivalents and serve as a mechanism to restore the cellular redox balance when cells experience reductive stress. These data suggest that killing of BCG by GSH may result from reductive stress that cannot be controlled. NRP BCG appears to be resistant to GSH-induced reductive stress.
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Affiliation(s)
- N D Patel
- Department of Biology, Pace University, 1 Pace Plaza, NY, NY 10038, United States
| | - R Lawrence
- Department of Biology, Pace University, 1 Pace Plaza, NY, NY 10038, United States
| | - M A Peteroy-Kelly
- Department of Biology, Pace University, 1 Pace Plaza, NY, NY 10038, United States.
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21
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Nývltová E, Smutná T, Tachezy J, Hrdý I. OsmC and incomplete glycine decarboxylase complex mediate reductive detoxification of peroxides in hydrogenosomes of Trichomonas vaginalis. Mol Biochem Parasitol 2016; 206:29-38. [PMID: 26794804 DOI: 10.1016/j.molbiopara.2016.01.006] [Citation(s) in RCA: 21] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/04/2015] [Revised: 01/05/2016] [Accepted: 01/12/2016] [Indexed: 01/30/2023]
Abstract
Osmotically inducible protein (OsmC) and organic hydroperoxide resistance protein (Ohr) are small, thiol-dependent peroxidases that comprise a family of prokaryotic protective proteins central to the defense against deleterious effects of organic hydroperoxides, which are reactive molecules that are formed during interactions between the host immune system and pathogens. Trichomonas vaginalis, a sexually transmitted parasite of humans, possesses OsmC homologues in its hydrogenosomes, anaerobic mitochondrial organelles that harbor enzymes and pathways that are sensitive to oxidative damage. The glycine decarboxylase complex (GDC), which consists of four proteins (i.e., L, H, P and T), is in eukaryotes exclusively mitochondrial enzymatic system that catalyzes oxidative decarboxylation and deamination of glycine. However, trichomonad hydrogenosomes contain only the L and H proteins, whose physiological functions are unknown. Here, we found that the hydrogenosomal L and H proteins constitute a lipoate-dependent redox system that delivers electrons from reduced nicotinamide adenine dinucleotide (NADH) to OsmC for the reductive detoxification of peroxides. Our searches of genome databases revealed that, in addition to prokaryotes, homologues of OsmC/Ohr family proteins with predicted mitochondrial localization are present in various eukaryotic lineages. Therefore, we propose that the novel OsmC-GDC-based redox system may not be limited to T. vaginalis.
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Affiliation(s)
- Eva Nývltová
- Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic
| | - Tamara Smutná
- Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic
| | - Jan Tachezy
- Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic
| | - Ivan Hrdý
- Department of Parasitology, Charles University in Prague, Faculty of Science, Prague, Czech Republic.
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22
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Abstract
The world health organization currently recognizes diarrhoeal diseases as a significant cause of death in children globally. Protozoan parasites such as Giardia and Entamoeba that thrive in the oxygen-deprived environment of the human gut are common etiological agents of diarrhoea. In the urogenital tract of humans, the anaerobic protozoan parasite Trichomonas vaginalis is notorious as the most common non-viral, sexually transmitted pathogen. Even with high medical impact, our understanding of anaerobic parasite physiology is scarce and as a result, treatment choices are limited. Fluorescent proteins (FPs) are invaluable tools as genetically encoded protein tags for advancing knowledge of cellular function. These FP tags emit fluorescent colours and once attached to a protein of interest, allow tracking of parasite proteins in the dynamic cellular space. Application of green FPs-like FPs in anaerobic protozoans is hindered by their oxygen dependency. In this review, we examine aspects of anaerobic parasite biology that clash with physio-chemical properties of FPs and limit their use as live-parasite protein tags. We expose novel FPs, such as miniSOG that do not require oxygen for signal production. The potential use of novel FPs has the opportunity to leverage the anaerobe parasitologist toolkit to that of aerobe parasitologist.
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23
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Vieira PDB, Silva NLF, Kist LW, Oliveira GMTD, Bogo MR, Carli GAD, Macedo AJ, Tasca T. Iron from haemoglobin and haemin modulates nucleotide hydrolysis in Trichomonas vaginalis. Mem Inst Oswaldo Cruz 2015; 110:201-8. [PMID: 25946243 PMCID: PMC4489450 DOI: 10.1590/0074-02760140320] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/01/2014] [Accepted: 01/26/2015] [Indexed: 12/14/2022] Open
Abstract
Extracellular ATP may act as a danger signalling molecule, inducing inflammation and
immune responses in infection sites. The ectonucleotidases NTPDase and
ecto-5’-nucleotidase are enzymes that modulate extracellular nucleotide levels; these
enzymes have been previously characterised in Trichomonas vaginalis.
Iron plays an important role in the complex trichomonal pathogenesis. Herein, the
effects of iron on growth, nucleotide hydrolysis and NTPDase gene expression in
T. vaginalis isolates from female and male patients were
evaluated. Iron from different sources sustained T. vaginalis
growth. Importantly, iron from haemoglobin (HB) and haemin (HM) enhanced NTPDase
activity in isolates from female patients and conversely reduced the enzyme activity
in isolates from male patients. Iron treatments could not alter the NTPDase
transcript levels in T. vaginalis. Furthermore, our results reveal a
distinct ATP, ADP and AMP hydrolysis profile between isolates from female and male
patients influenced by iron from HB and HM. Our data indicate the participation of
NTPDase and ecto-5’-nucleotidase in the establishment of trichomonas infection
through ATP degradation and adenosine production influenced by iron.
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Affiliation(s)
| | | | - Luiza Wilges Kist
- Laboratório de Biologia Genômica e Molecular, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, RS, Brasil
| | | | - Maurício Reis Bogo
- Laboratório de Biologia Genômica e Molecular, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, RS, Brasil
| | - Geraldo Atillio de Carli
- Instituto de Geriatria e Gerontologia, Pontifícia Universidade Católica do Rio Grande do Sul, Porto Alegre, RS, Brasil
| | - Alexandre José Macedo
- Faculdade de Farmácia, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brasil
| | - Tiana Tasca
- Faculdade de Farmácia, Universidade Federal do Rio Grande do Sul, Porto Alegre, RS, Brasil
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24
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Kutsumura N, Nakajima R, Koyama Y, Miyata Y, Saitoh T, Yamamoto N, Iwata S, Fujii H, Nagase H. Investigation of 7-benzylidenenaltrexone derivatives as a novel structural antitrichomonal lead compound. Bioorg Med Chem Lett 2015; 25:4890-4892. [DOI: 10.1016/j.bmcl.2015.06.002] [Citation(s) in RCA: 5] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/28/2015] [Revised: 05/28/2015] [Accepted: 06/02/2015] [Indexed: 12/13/2022]
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Korosh T, Jordan KD, Wu JS, Yarlett N, Upmacis RK. Eicosapentaenoic Acid Modulates Trichomonas vaginalis Activity. J Eukaryot Microbiol 2015; 63:153-61. [PMID: 26333486 DOI: 10.1111/jeu.12263] [Citation(s) in RCA: 3] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/16/2015] [Accepted: 08/26/2015] [Indexed: 01/23/2023]
Abstract
Trichomonas vaginalis is a sexually transmitted parasite and, while it is often asymptomatic in males, the parasite is associated with disease in both sexes. Metronidazole is an effective treatment for trichomoniasis, but resistant strains have evolved and, thus, it has become necessary to investigate other possible therapies. In this study, we examined the effects of native and oxidized forms of the sodium salts of eicosapentaenoic, docosahexaenoic, and arachidonic acids on T. vaginalis activity. Eicosapentaenoic acid was the most toxic with 190 and 380 μM causing approximately 90% cell death in Casu2 and ATCC 50142 strains, respectively. In contrast, oxidized eicosapentaenoic acid was the least toxic, requiring > 3 mM to inhibit activity, while low levels (10 μM) were associated with increased parasite density. Mass spectrometric analysis of oxidized eicosapentaenoic acid revealed C20 products containing one to six additional oxygen atoms and various degrees of bond saturation. These results indicate that eicosapentaenoic acid has different effects on T. vaginalis survival, depending on whether it is present in the native or oxidized form. A better understanding of lipid metabolism in T. vaginalis may facilitate the design of synthetic fatty acids that are effective for the treatment of metronidazole-resistant T. vaginalis.
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Affiliation(s)
- Travis Korosh
- The Haskins Laboratories, Department of Chemistry and Physical Sciences, Pace University, 41 Park Row, New York City, New York, 10038
| | - Kelsey D Jordan
- The Haskins Laboratories, Department of Chemistry and Physical Sciences, Pace University, 41 Park Row, New York City, New York, 10038
| | - Ja-Shin Wu
- The Haskins Laboratories, Department of Chemistry and Physical Sciences, Pace University, 41 Park Row, New York City, New York, 10038
| | - Nigel Yarlett
- The Haskins Laboratories, Department of Chemistry and Physical Sciences, Pace University, 41 Park Row, New York City, New York, 10038
| | - Rita K Upmacis
- The Haskins Laboratories, Department of Chemistry and Physical Sciences, Pace University, 41 Park Row, New York City, New York, 10038
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26
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Elvitigala DAS, Whang I, Lee J. Molecular profiling and functional insights of rock bream (Oplegnathus fasciatus) thioredoxin reductase 3-like molecule: investigation of its transcriptional modulation in response to live pathogen stress. Gene 2015; 570:122-31. [PMID: 26055087 DOI: 10.1016/j.gene.2015.06.007] [Citation(s) in RCA: 1] [Impact Index Per Article: 0.1] [Reference Citation Analysis] [Abstract] [Key Words] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/03/2015] [Revised: 05/20/2015] [Accepted: 06/03/2015] [Indexed: 01/22/2023]
Abstract
The thioredoxin (Trx) system plays a significant role in cellular antioxidative defense by dismutating the surpluses of reactive oxygen species. Thus, the role of thioredoxin reductase (TrxR) cannot be ignored, owing to its participation in initiating the Trx enzyme cascade. Here, we report the identification and molecular characterization of a teleostean TrxR (RbTrxR-3) ortholog that showed high similarity with the TrxR-3 isoforms of other vertebrates. The complete RbTrxR-3 coding sequence comprised 1800 nucleotides, encoding a 600-amino acid protein with a predicted molecular mass of ~66 kDa. RbTrxR-3 consisted of 16 exons separated by 15 introns and had a total length of 12,658 bp. In silico analysis of the RbTrxR-3 protein sequence revealed that it possesses typical TrxR domain architecture. Moreover, using multiple sequence alignment and pairwise sequence alignment strategies, we showed that RbTrxR-3 has high overall sequence similarity to other teleostean TrxR-3 proteins, including highly conserved active site residues. Phylogenetic reconstruction of RbTrxR-3 affirmed its close evolutionary relationship with fish TrxR-3 orthologs, as indicated by its clustering pattern. RbTrxR-3 transcriptional analysis, performed using quantitative polymerase chain reaction (qPCR), showed that RbTrxR-3 was ubiquitously distributed, with the highest level of mRNA expression in the blood, followed by the gill, and liver. Live bacterial and viral stimuli triggered the modulation of RbTrxR-3 basal transcription in liver tissues that correlated temporally with that of its putative substrate, rock bream thioredoxin1 under the same conditions of pathogenic stress. Finally, resembling the typical function of TrxR protein, purified recombinant RbTrxR-3 showed detectable dose-dependent thiol reductase activity against 5,5'-dithiobis (2-nitrobenzoic) acid. Taken together, these results suggest that RbTrxR-3 plays a role in the host Trx system under conditions of oxidative and pathogenic stress.
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Affiliation(s)
- Don Anushka Sandaruwan Elvitigala
- Department of Marine Life Sciences, School of Marine Biomedical Sciences, Jeju National University, Jeju Self-Governing Province 690-756, Republic of Korea; Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province 690-756, Republic of Korea
| | - Ilson Whang
- Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province 690-756, Republic of Korea.
| | - Jehee Lee
- Department of Marine Life Sciences, School of Marine Biomedical Sciences, Jeju National University, Jeju Self-Governing Province 690-756, Republic of Korea; Fish Vaccine Research Center, Jeju National University, Jeju Self-Governing Province 690-756, Republic of Korea.
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27
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Antioxidant defences of Spironucleus vortens: Glutathione is the major non-protein thiol. Mol Biochem Parasitol 2014; 196:45-52. [DOI: 10.1016/j.molbiopara.2014.07.010] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/02/2013] [Revised: 07/21/2014] [Accepted: 07/28/2014] [Indexed: 11/17/2022]
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28
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Siavoshi F, Saniee P. Vacuoles of Candida yeast as a specialized niche for Helicobacter pylori. World J Gastroenterol 2014; 20:5263-5273. [PMID: 24833856 PMCID: PMC4017041 DOI: 10.3748/wjg.v20.i18.5263] [Citation(s) in RCA: 43] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 10/15/2013] [Revised: 01/09/2014] [Accepted: 02/26/2014] [Indexed: 02/06/2023] Open
Abstract
Helicobacter pylori (H. pylori) are resistant to hostile gastric environments and antibiotic therapy, reflecting the possibility that they are protected by an ecological niche, such as inside the vacuoles of human epithelial and immune cells. Candida yeast may also provide such an alternative niche, as fluorescently labeled H. pylori were observed as fast-moving and viable bacterium-like bodies inside the vacuoles of gastric, oral, vaginal and foodborne Candida yeasts. In addition, H. pylori-specific genes and proteins were detected in samples extracted from these yeasts. The H. pylori present within these yeasts produce peroxiredoxin and thiol peroxidase, providing the ability to detoxify oxygen metabolites formed in immune cells. Furthermore, these bacteria produce urease and VacA, two virulence determinants of H. pylori that influence phago-lysosome fusion and bacterial survival in macrophages. Microscopic observations of H. pylori cells in new generations of yeasts along with amplification of H. pylori-specific genes from consecutive generations indicate that new yeasts can inherit the intracellular H. pylori as part of their vacuolar content. Accordingly, it is proposed that yeast vacuoles serve as a sophisticated niche that protects H. pylori against the environmental stresses and provides essential nutrients, including ergosterol, for its growth and multiplication. This intracellular establishment inside the yeast vacuole likely occurred long ago, leading to the adaptation of H. pylori to persist in phagocytic cells. The presence of these bacteria within yeasts, including foodborne yeasts, along with the vertical transmission of yeasts from mother to neonate, provide explanations for the persistence and propagation of H. pylori in the human population. This Topic Highlight reviews and discusses recent evidence regarding the evolutionary adaptation of H. pylori to thrive in host cell vacuoles.
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Novel functions of an iron-sulfur flavoprotein from Trichomonas vaginalis hydrogenosomes. Antimicrob Agents Chemother 2014; 58:3224-32. [PMID: 24663020 DOI: 10.1128/aac.02320-13] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.5] [Reference Citation Analysis] [Abstract] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/10/2023] Open
Abstract
Iron-sulfur flavoproteins (Isf) are flavin mononucleotide (FMN)- and FeS cluster-containing proteins commonly encountered in anaerobic prokaryotes. However, with the exception of Isf from Methanosarcina thermophila, which participates in oxidative stress management by removing oxygen and hydrogen peroxide, none of these proteins has been characterized in terms of function. Trichomonas vaginalis, a sexually transmitted eukaryotic parasite of humans, was found to express several iron-sulfur flavoprotein (TvIsf) homologs in its hydrogenosomes. We show here that in addition to having oxygen-reducing activity, the recombinant TvIsf also functions as a detoxifying reductase of metronidazole and chloramphenicol, both of which are antibiotics effective against a variety of anaerobic microbes. TvIsf can utilize both NADH and reduced ferredoxin as electron donors. Given the prevalence of Isf in anaerobic prokaryotes, we propose that these proteins are central to a novel defense mechanism against xenobiotics.
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Molecular dynamic simulation and inhibitor prediction of cysteine synthase structured model as a potential drug target for trichomoniasis. BIOMED RESEARCH INTERNATIONAL 2013; 2013:390920. [PMID: 24073401 PMCID: PMC3773994 DOI: 10.1155/2013/390920] [Citation(s) in RCA: 7] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/26/2013] [Accepted: 07/14/2013] [Indexed: 12/03/2022]
Abstract
In our presented research, we made an attempt to predict the 3D model for cysteine synthase (A2GMG5_TRIVA) using homology-modeling approaches. To investigate deeper into the predicted structure, we further performed a molecular dynamics simulation for 10 ns and calculated several supporting analysis for structural properties such as RMSF, radius of gyration, and the total energy calculation to support the predicted structured model of cysteine synthase. The present findings led us to conclude that the proposed model is stereochemically stable. The overall PROCHECK G factor for the homology-modeled structure was −0.04. On the basis of the virtual screening for cysteine synthase against the NCI subset II molecule, we present the molecule 1-N, 4-N-bis [3-(1H-benzimidazol-2-yl) phenyl] benzene-1,4-dicarboxamide (ZINC01690699) having the minimum energy score (−13.0 Kcal/Mol) and a log P value of 6 as a potential inhibitory molecule used to inhibit the growth of T. vaginalis infection.
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Gould SB, Woehle C, Kusdian G, Landan G, Tachezy J, Zimorski V, Martin WF. Deep sequencing of Trichomonas vaginalis during the early infection of vaginal epithelial cells and amoeboid transition. Int J Parasitol 2013; 43:707-19. [DOI: 10.1016/j.ijpara.2013.04.002] [Citation(s) in RCA: 53] [Impact Index Per Article: 4.4] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/04/2012] [Revised: 04/08/2013] [Accepted: 04/09/2013] [Indexed: 01/15/2023]
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Gretes MC, Poole LB, Karplus PA. Peroxiredoxins in parasites. Antioxid Redox Signal 2012; 17:608-33. [PMID: 22098136 PMCID: PMC3373223 DOI: 10.1089/ars.2011.4404] [Citation(s) in RCA: 70] [Impact Index Per Article: 5.4] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/14/2011] [Accepted: 11/18/2011] [Indexed: 12/11/2022]
Abstract
SIGNIFICANCE Parasite survival and virulence relies on effective defenses against reactive oxygen and nitrogen species produced by the host immune system. Peroxiredoxins (Prxs) are ubiquitous enzymes now thought to be central to such defenses and, as such, have potential value as drug targets and vaccine antigens. RECENT ADVANCES Plasmodial and kinetoplastid Prx systems are the most extensively studied, yet remain inadequately understood. For many other parasites our knowledge is even less well developed. Through parasite genome sequencing efforts, however, the key players are being discovered and characterized. Here we describe what is known about the biochemistry, regulation, and cell biology of Prxs in parasitic protozoa, helminths, and fungi. At least one Prx is found in each parasite with a sequenced genome, and a notable theme is the common patterns of expression, localization, and functionality among sequence-similar Prxs in related species. CRITICAL ISSUES The nomenclature of Prxs from parasites is in a state of disarray, causing confusion and making comparative inferences difficult. Here we introduce a systematic Prx naming convention that is consistent between organisms and informative about structural and evolutionary relationships. FUTURE DIRECTIONS The new nomenclature should stimulate the crossfertilization of ideas among parasitologists and with the broader redox research community. The diverse parasite developmental stages and host environments present complex systems in which to explore the variety of roles played by Prxs, with a view toward parlaying what is learned into novel therapies and vaccines that are urgently needed.
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Affiliation(s)
- Michael C. Gretes
- Department of Biochemistry & Biophysics, Oregon State University, Corvallis, Oregon
| | - Leslie B. Poole
- Department of Biochemistry, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - P. Andrew Karplus
- Department of Biochemistry & Biophysics, Oregon State University, Corvallis, Oregon
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Kay C, Woodward KD, Lawler K, Self TJ, Dyall SD, Kerr ID. The ATP-binding cassette proteins of the deep-branching protozoan parasite Trichomonas vaginalis. PLoS Negl Trop Dis 2012; 6:e1693. [PMID: 22724033 PMCID: PMC3378599 DOI: 10.1371/journal.pntd.0001693] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.5] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/11/2011] [Accepted: 03/18/2012] [Indexed: 02/06/2023] Open
Abstract
The ATP binding cassette (ABC) proteins are a family of membrane transporters and regulatory proteins responsible for diverse and critical cellular process in all organisms. To date, there has been no attempt to investigate this class of proteins in the infectious parasite Trichomonas vaginalis. We have utilized a combination of bioinformatics, gene sequence analysis, gene expression and confocal microscopy to investigate the ABC proteins of T. vaginalis. We demonstrate that, uniquely among eukaryotes, T. vaginalis possesses no intact full-length ABC transporters and has undergone a dramatic expansion of some ABC protein sub-families. Furthermore, we provide preliminary evidence that T. vaginalis is able to read through in-frame stop codons to express ABC transporter components from gene pairs in a head-to-tail orientation. Finally, with confocal microscopy we demonstrate the expression and endoplasmic reticulum localization of a number of T. vaginalis ABC transporters. The parasite Trichomonas vaginalis infects in excess of 100 million people per year, and is a contributory factor to enhanced transmission rates of HIV, the causative virus in AIDS. As such, T. vaginalis infection is an important public health concern. Understanding the biology of the organism is important to determine aspects of the response to drug treatment, host:parasite interactions and so on. We have investigated an important family of proteins – the ATP binding cassette transporters – which are present in the membranes of all cells, and which contribute to a diverse spectrum of important cellular processes. The ABC transporters of T. vaginalis were identified by analysis of primary amino acid sequence data, and examined by subsequent protein and gene expression studies. Our most important conclusion is that – uniquely amongst eukaryotes - T. vaginalis has no ABC transporters capable of acting as monomers. In other words, its ABC transporters must all act by forming functional complexes with other ABC proteins. This has implications for our understanding not just of the parasite's biology, but also its evolution. In summary our analysis opens up the path for future research of individual members of the ABC protein family in T. vaginalis.
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Affiliation(s)
- Christopher Kay
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
| | - Katharine D. Woodward
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
| | - Karen Lawler
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
| | - Tim J. Self
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
| | - Sabrina D. Dyall
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
- Department of Biosciences, University of Mauritius, Reduit, Mauritius
| | - Ian D. Kerr
- School of Biomedical Sciences, University of Nottingham Medical School, Queen's Medical Centre, Nottingham, United Kingdom
- * E-mail:
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Leitsch D, Drinić M, Kolarich D, Duchêne M. Down-regulation of flavin reductase and alcohol dehydrogenase-1 (ADH1) in metronidazole-resistant isolates of Trichomonas vaginalis. Mol Biochem Parasitol 2012; 183:177-83. [PMID: 22449940 PMCID: PMC3341570 DOI: 10.1016/j.molbiopara.2012.03.003] [Citation(s) in RCA: 30] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/08/2011] [Revised: 03/01/2012] [Accepted: 03/06/2012] [Indexed: 01/23/2023]
Abstract
The microaerophilic parasite Trichomonas vaginalis is a causative agent of painful vaginitis or urethritis, termed trichomoniasis, and can also cause preterm delivery or stillbirth. Treatment of trichomoniasis is almost exclusively based on the nitroimidazole drugs metronidazole and tinidazole. Metronidazole resistance in T. vaginalis does occur and is often associated with treatment failure. In most cases, metronidazole-resistant isolates remain susceptible to tinidazole, but cross resistance between the two closely related drugs can be a problem. In this study we measured activities of thioredoxin reductase and flavin reductase in four metronidazole-susceptible and five metronidazole-resistant isolates. These enzyme activities had been previously found to be downregulated in T. vaginalis with high-level metronidazole resistance induced in the laboratory. Further, we aimed at identifying factors causing metronidazole resistance and compared the protein expression profiles of all nine isolates by application of two-dimensional gel electrophoresis (2DE). Thioredoxin reductase activity was nearly equal in all strains assayed but flavin reductase activity was clearly down-regulated, or even absent, in metronidazole-resistant strains. Since flavin reductase has been shown to reduce oxygen to hydrogen peroxide, its down-regulation could significantly contribute to the impairment of oxygen scavenging as reported by others for metronidazole-resistant strains. Analysis by 2DE revealed down-regulation of alcohol dehydrogenase 1 (ADH1) in strains with reduced sensitivity to metronidazole, an enzyme that could be involved in detoxification of intracellular acetaldehyde.
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Affiliation(s)
- David Leitsch
- Institute of Specific Prophylaxis and Tropical Medicine at the Center for Pathophysiology, Infectiology and Immunology, Medical University of Vienna, 1090 Vienna, Austria.
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Kappes B, Tews I, Binter A, Macheroux P. PLP-dependent enzymes as potential drug targets for protozoan diseases. BIOCHIMICA ET BIOPHYSICA ACTA 2011; 1814:1567-76. [PMID: 21884827 DOI: 10.1016/j.bbapap.2011.07.018] [Citation(s) in RCA: 27] [Impact Index Per Article: 1.9] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Subscribe] [Scholar Register] [Received: 02/13/2011] [Revised: 07/01/2011] [Accepted: 07/18/2011] [Indexed: 11/20/2022]
Abstract
The chemical properties of the B(6) vitamers are uniquely suited for wide use as cofactors in essential reactions, such as decarboxylations and transaminations. This review addresses current efforts to explore vitamin B(6) dependent enzymatic reactions as drug targets. Several current targets are described that are found amongst these enzymes. The focus is set on diseases caused by protozoan parasites. Comparison across a range of these organisms allows insight into the distribution of potential targets, many of which may be of interest in the development of broad range anti-protozoan drugs. This article is part of a Special Issue entitled: Pyridoxal Phosphate Enzymology.
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Affiliation(s)
- Barbara Kappes
- University Hospital Heidelberg, Department of Infectious Diseases, Parasitology, Im Neuenheimer Feld 324, 69120 Heidelberg, Germany
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Characterization of Deinococcus radiophilus thioredoxin reductase active with both NADH and NADPH. J Microbiol 2010; 48:637-43. [PMID: 21046342 DOI: 10.1007/s12275-010-0283-7] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2010] [Accepted: 10/08/2010] [Indexed: 09/29/2022]
Abstract
Thioredoxin reductase (TrxR, EC 1.6.4.5) of Deinococcus radiophilus was purified by steps of sonication, ammonium sulfate fractionation, 2'5' ADP Sepharose 4B affinity chromatography, and Sephadex G-100 gel filtration. The purified TrxR, which was active with both NADPH and NADH, gave a 368 U/mg protein of specific activity with 478-fold purification and 18% recovery from the cell-free extract. An isoelectric point of the purified enzymes was ca. 4.5. The molecular weights of the purified TrxR estimated by PAGE and gel filtration were about 63.1 and 72.2 kDa, respectively. The molecular mass of a TrxR subunit is 37 kDa. This suggests that TrxR definitely belongs to low molecular weight TrxR (L-TrxR). The Km and Vmax of TrxR for NADPH are 12.5 μM and 25 μM/min, whereas those for NADH are 30.2 μM and 192 μM/min. The Km and Vmax for 5, 5'-dithio-bis-2-nitrobenzoic acid (DTNB, a substituted substrate for thioredoxin) are 463 μM and 756 μM/min, respectively. The presence of FAD in TrxR was confirmed with the absorbance peaks at 385 and 460 nm. The purified TrxR was quite stable from pH 3 to 9, and was thermo-stable up to 70°C. TrxR activity was drastically reduced (ca. 70%) by Cu(2+), Zn(2+), Hg(2+), and Cd(2+), but moderately reduced (ca. 50%) by Ag(+). A significant inhibition of TrxR by N-ethylmaleimide suggests an occurrence of cysteine at its active sites. Amino acid sequences at the N-terminus of purified TrxR are H(2)N-Ser-Glu-Gln-Ala-Gln-Met-Tyr-Asp-Val-Ile-Ile-Val-Gly-Gly-Gly-Pro-Ala-Gly-Leu-Thr-Ala-COOH. These sequences show high similarity with TrxRs reported in Archaea, such as Methanosarcina mazei, Archaeoglobus fulgidus etc.
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Westrop GD, Georg I, Coombs GH. The mercaptopyruvate sulfurtransferase of Trichomonas vaginalis links cysteine catabolism to the production of thioredoxin persulfide. J Biol Chem 2009; 284:33485-94. [PMID: 19762467 DOI: 10.1074/jbc.m109.054320] [Citation(s) in RCA: 45] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Trichomonas vaginalis is a protozoan parasite of humans that is able to synthesize cysteine de novo using cysteine synthase but does not produce glutathione. In this study, high pressure liquid chromatography analysis confirmed that cysteine is the major intracellular redox buffer by showing that T. vaginalis contains high levels of cysteine ( approximately 600 mum) comprising more than 70% of the total thiols detected. To investigate possible mechanisms for the regulation of cysteine levels in T. vaginalis, we have characterized enzymes of the mercaptopyruvate pathway. This consists of an aspartate aminotransferase (TvAspAT1), which transaminates cysteine to form 3-mercaptopyruvate (3-MP), and mercaptopyruvate sulfurtransferase (TvMST), which transfers the sulfur of 3-MP to a nucleophilic acceptor, generating pyruvate. TvMST has high activity with 3-MP as a sulfur donor and can use several thiol compounds as sulfur acceptor substrates. Our analysis indicated that TvMST has a k(cat)/K(m) for reduced thioredoxin of 6.2 x 10(7) m(-1) s(-1), more than 100-fold higher than that observed for beta-mercaptoethanol and cysteine, suggesting that thioredoxin is a preferred substrate for TvMST. Thiol trapping and mass spectrometry provided direct evidence for the formation of thioredoxin persulfide as a product of this reaction. The thioredoxin persulfide could serve a biological function such as the transfer of the persulfide to a target protein or the sequestered release of sulfide for biosynthesis. Changes in MST activity of T. vaginalis in response to variation in the supply of exogenous cysteine are suggestive of a role for the mercaptopyruvate pathway in the removal of excess intracellular cysteine, redox homeostasis, and antioxidant defense.
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Affiliation(s)
- Gareth D Westrop
- Strathclyde Institute of Pharmacy and Biomedical Sciences, Univesity of Strathclyde, Glasgow G4 0NR, United Kingdom.
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Gehrig S, Efferth T. Development of Drug Resistance in Trichomonas vaginalis and its Overcoming with Natural Products. ACTA ACUST UNITED AC 2009. [DOI: 10.2174/1874847300902010021] [Citation(s) in RCA: 12] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Trichomoniasis is an infectious disease afflicting women worldwide. The protozoan parasite Trichomonas vaginalis is the causative agent of this sexually-transmitted disease, including also men in its infection cycle. The disease is usually not life-threatening, but has been associated with the development of cervical cancer and increased susceptibility to HIV. Approved drugs are 5-nitroimidazoles, with metronidazole being the drug of first choice. These drugs act via induction of oxidative stress and DNA-damage, leading to cell death in the parasite. Nevertheless, with the development of resistant T. vaginalis strains the treatment of the disease becomes exceedingly difficult. Mechanisms of drug resistance are characterized by reduced expression or even loss of proteins necessary for drug activation and a decreased reductive nature in the parasite. A promising strategy for research into new drugs and moreover, to overcome drug resistance, are compounds derived from natural sources. The present study provides a summary of all so far investigated small molecules with antitrichomonal activity; promisingly, some also show efficacy against resistant strains. Whereas the list of chemically characterized compounds derived from plants is rather short, literature provides immense applications of crude plant extracts tested against T. vaginalis. This demonstrates the absence of studies in this field aimed to identify and isolate single natural products exhibiting antitrichomonal features. Likewise, elucidating their mode of action on a molecular basis is of paramount importance
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Leitsch D, Kolarich D, Binder M, Stadlmann J, Altmann F, Duchêne M. Trichomonas vaginalis: metronidazole and other nitroimidazole drugs are reduced by the flavin enzyme thioredoxin reductase and disrupt the cellular redox system. Implications for nitroimidazole toxicity and resistance. Mol Microbiol 2009; 72:518-36. [PMID: 19415801 DOI: 10.1111/j.1365-2958.2009.06675.x] [Citation(s) in RCA: 106] [Impact Index Per Article: 6.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/18/2022]
Abstract
Infections with the microaerophilic parasite Trichomonas vaginalis are treated with the 5-nitroimidazole drug metronidazole, which is also in use against Entamoeba histolytica, Giardia intestinalis and microaerophilic/anaerobic bacteria. Here we report that in T. vaginalis the flavin enzyme thioredoxin reductase displays nitroreductase activity with nitroimidazoles, including metronidazole, and with the nitrofuran drug furazolidone. Reactive metabolites of metronidazole and other nitroimidazoles form covalent adducts with several proteins that are known or assumed to be associated with thioredoxin-mediated redox regulation, including thioredoxin reductase itself, ribonucleotide reductase, thioredoxin peroxidase and cytosolic malate dehydrogenase. Disulphide reducing activity of thioredoxin reductase was greatly diminished in extracts of metronidazole-treated cells and intracellular non-protein thiol levels were sharply decreased. We generated a highly metronidazole-resistant cell line that displayed only minimal thioredoxin reductase activity, not due to diminished expression of the enzyme but due to the lack of its FAD cofactor. Reduction of free flavins, readily observed in metronidazole-susceptible cells, was also absent in the resistant cells. On the other hand, iron-depleted T. vaginalis cells, expressing only minimal amounts of PFOR and hydrogenosomal malate dehydrogenase, remained fully susceptible to metronidazole. Thus, taken together, our data suggest a flavin-based mechanism of metronidazole activation and thereby challenge the current model of hydrogenosomal activation of nitroimidazole drugs.
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Affiliation(s)
- David Leitsch
- Department of Specific Prophylaxis and Tropical Medicine at the Center for Physiology, Pathophysiology and Immunology, Medical University of Vienna, Vienna, Austria.
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BILIC I, LEBERL M, HESS M. Identification and molecular characterization of numerous Histomonas meleagridis proteins using a cDNA library. Parasitology 2009; 136:379-91. [PMID: 19154645 PMCID: PMC2957082 DOI: 10.1017/s0031182008005477] [Citation(s) in RCA: 15] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022]
Abstract
SUMMARYHistomonas meleagridis is a protozoan parasite of various galliform birds causing a type of enterohepatitis termed histomonosis or 'blackhead disease'. Due to the ban of chemotherapeutic substances and an increase in free-range poultry production, histomonosis is currently a re-emerging disease. So far limited molecular knowledge is available. In the present work, mRNAs coding for antigenic proteins of H. meleagridis were identified. For this purpose, a cDNA expression library was constructed from a mono-eukaryotic culture of H. meleagridis. The library was screened with polyclonal rabbit serum raised against purified H. meleagridis trophozoites. Polyclonal rabbit serum specifically recognized the same major H. meleagridis antigens as chicken and turkey sera originating from animal trials, but displayed a significantly lower bacteria-dependent background signal. After 2 rounds of screening, a total of 95 positive clones were sequenced. Bioinformatics analyses were performed on nucleotide and deduced amino acid sequences, identifying 37 unique clones. Based on the homology to other protozoan parasites, mostly Trichomonas vaginalis, the clones were grouped according to functional aspects: structural proteins, possible surface proteins, oxygen reducing proteins, ribosomal proteins, protein kinases and various other intracellular proteins.
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Affiliation(s)
- I. BILIC
- Clinic for Avian, Reptile and Fish Medicine, University of Veterinary Medicine, Vienna, Veterinaerplatz 1, 1210 Vienna, Austria
| | - M. LEBERL
- Clinic for Avian, Reptile and Fish Medicine, University of Veterinary Medicine, Vienna, Veterinaerplatz 1, 1210 Vienna, Austria
| | - M. HESS
- Clinic for Avian, Reptile and Fish Medicine, University of Veterinary Medicine, Vienna, Veterinaerplatz 1, 1210 Vienna, Austria
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Flavodiiron protein from Trichomonas vaginalis hydrogenosomes: the terminal oxygen reductase. EUKARYOTIC CELL 2008; 8:47-55. [PMID: 19011120 DOI: 10.1128/ec.00276-08] [Citation(s) in RCA: 55] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
Trichomonas vaginalis is one of a few eukaryotes that have been found to encode several homologues of flavodiiron proteins (FDPs). Widespread among anaerobic prokaryotes, these proteins are believed to function as oxygen and/or nitric oxide reductases to provide protection against oxidative/nitrosative stresses and host immune responses. One of the T. vaginalis FDP homologues is equipped with a hydrogenosomal targeting sequence and is expressed in the hydrogenosomes, oxygen-sensitive organelles that participate in carbohydrate metabolism and assemble iron-sulfur clusters. The bacterial homologues characterized thus far have been dimers or tetramers; the trichomonad protein is a dimer of identical 45-kDa subunits, each noncovalently binding one flavin mononucleotide. The protein reduces dioxygen to water but is unable to utilize nitric oxide as a substrate, similarly to its closest homologue from another human parasite Giardia intestinalis and related archaebacterial proteins. T. vaginalis FDP is able to accept electrons derived from pyruvate or NADH via ferredoxin and is proposed to play a role in the protection of hydrogenosomes against oxygen.
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Protein import into hydrogenosomes of Trichomonas vaginalis involves both N-terminal and internal targeting signals: a case study of thioredoxin reductases. EUKARYOTIC CELL 2008; 7:1750-7. [PMID: 18676956 DOI: 10.1128/ec.00206-08] [Citation(s) in RCA: 39] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 11/20/2022]
Abstract
The parabasalian flagellate Trichomonas vaginalis harbors mitochondrion-related and H(2)-producing organelles of anaerobic ATP synthesis, called hydrogenosomes, which harbor oxygen-sensitive enzymes essential to its pyruvate metabolism. In the human urogenital tract, however, T. vaginalis is regularly exposed to low oxygen concentrations and therefore must possess antioxidant systems protecting the organellar environment against the detrimental effects of molecular oxygen and reactive oxygen species. We have identified two closely related hydrogenosomal thioredoxin reductases (TrxRs), the hitherto-missing component of a thioredoxin-linked hydrogenosomal antioxidant system. One of the two hydrogenosomal TrxR isoforms, TrxRh1, carried an N-terminal extension resembling known hydrogenosomal targeting signals. Expression of hemagglutinin-tagged TrxRh1 in transfected T. vaginalis cells revealed that its N-terminal extension was necessary to import the protein into the organelles. The second hydrogenosomal TrxR isoform, TrxRh2, had no N-terminal targeting signal but was nonetheless efficiently targeted to hydrogenosomes. N-terminal presequences from hydrogenosomal proteins with known processing sites, i.e., the alpha subunit of succinyl coenzyme A synthetase (SCSalpha) and pyruvate:ferredoxin oxidoreductase A, were investigated for their ability to direct mature TrxRh1 to hydrogenosomes. Neither presequence directed TrxRh1 to hydrogenosomes, indicating that neither extension is, by itself, sufficient for hydrogenosomal targeting. Moreover, SCSalpha lacking its N-terminal extension was efficiently imported into hydrogenosomes, indicating that this extension is not required for import of this major hydrogenosomal protein. The finding that some hydrogenosomal enzymes require N-terminal signals for import but that in others the N-terminal extension is not necessary for targeting indicates the presence of additional targeting signals within the mature subunits of several hydrogenosome-localized proteins.
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Leitsch D, Kolarich D, Wilson IBH, Altmann F, Duchêne M. Nitroimidazole action in Entamoeba histolytica: a central role for thioredoxin reductase. PLoS Biol 2008; 5:e211. [PMID: 17676992 PMCID: PMC1933457 DOI: 10.1371/journal.pbio.0050211] [Citation(s) in RCA: 119] [Impact Index Per Article: 7.0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/30/2007] [Accepted: 06/07/2007] [Indexed: 02/06/2023] Open
Abstract
Metronidazole, a 5-nitroimidazole drug, has been the gold standard for several decades in the treatment of infections with microaerophilic protist parasites, including Entamoeba histolytica. For activation, the drug must be chemically reduced, but little is known about the targets of the active metabolites. Applying two-dimensional gel electrophoresis and mass spectrometry, we searched for protein targets in E. histolytica. Of all proteins visualized, only five were found to form adducts with metronidazole metabolites: thioredoxin, thioredoxin reductase, superoxide dismutase, purine nucleoside phosphorylase, and a previously unknown protein. Recombinant thioredoxin reductase carrying the modification displayed reduced enzymatic activity. In treated cells, essential non-protein thiols such as free cysteine were also affected by covalent adduct formation, their levels being drastically reduced. Accordingly, addition of cysteine allowed E. histolytica to survive in the presence of otherwise lethal metronidazole concentrations and reduced protein adduct formation. Finally, we discovered that thioredoxin reductase reduces metronidazole and other nitro compounds, suggesting a new model of metronidazole activation in E. histolytica with a central role for thioredoxin reductase. By reducing metronidazole, the enzyme renders itself and associated thiol-containing proteins vulnerable to adduct formation. Because thioredoxin reductase is a ubiquitous enzyme, similar processes could occur in other eukaryotic or prokaryotic organisms. The protist parasites Entamoeba histolytica, Trichomonas vaginalis, and Giardia intestinalis grow in environments with low oxygen concentration. Infections with these parasites are commonly treated with metronidazole, a nitroimidazole drug that must be reduced for activation, resulting in several toxic metabolites. We examined the soluble proteome of metronidazole-treated E. histolytica cells for target proteins of these metabolites, applying two-dimensional gel electrophoresis and mass spectrometry. Of about 1,500 proteins visualized, only five formed covalent adducts with metronidazole metabolites, including thioredoxin, thioredoxin reductase, and superoxide dismutase. Metronidazole-bound thioredoxin reductase displayed diminished activity. In addition to these proteins, small thiol molecules, including cysteine, formed adducts with metronidazole. Supplementation with cysteine allowed the cells to survive otherwise lethal metronidazole concentrations. Finally, we discovered that one of the modified proteins, thioredoxin reductase, reduces metronidazole, suggesting a central role for this enzyme with regard to metronidazole toxicity. Taken together, our work reveals a new area of molecular interactions of activated metronidazole with cellular components. Because thioredoxin reductase is a ubiquitous enzyme, similar processes could also occur in other eukaryotic or prokaryotic organisms. Metronidazole is used for treatment of infections with microaerophilic protist parasites. Here, a new model of metronidazole activation is proposed, with a central role for thioredoxin reductase.
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Affiliation(s)
- David Leitsch
- Department of Specific Prophylaxis and Tropical Medicine, Center for Physiology and Pathophysiology, Medical University of Vienna, Vienna, Austria
| | - Daniel Kolarich
- Department of Chemistry, University of Natural Resources and Applied Life Sciences, Vienna, Austria
| | - Iain B. H Wilson
- Department of Chemistry, University of Natural Resources and Applied Life Sciences, Vienna, Austria
| | - Friedrich Altmann
- Department of Chemistry, University of Natural Resources and Applied Life Sciences, Vienna, Austria
| | - Michael Duchêne
- Department of Specific Prophylaxis and Tropical Medicine, Center for Physiology and Pathophysiology, Medical University of Vienna, Vienna, Austria
- * To whom correspondence should be addressed. E-mail:
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Ali V, Nozaki T. Current therapeutics, their problems, and sulfur-containing-amino-acid metabolism as a novel target against infections by "amitochondriate" protozoan parasites. Clin Microbiol Rev 2007; 20:164-87. [PMID: 17223627 PMCID: PMC1797636 DOI: 10.1128/cmr.00019-06] [Citation(s) in RCA: 127] [Impact Index Per Article: 7.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/30/2023] Open
Abstract
The "amitochondriate" protozoan parasites of humans Entamoeba histolytica, Giardia intestinalis, and Trichomonas vaginalis share many biochemical features, e.g., energy and amino acid metabolism, a spectrum of drugs for their treatment, and the occurrence of drug resistance. These parasites possess metabolic pathways that are divergent from those of their mammalian hosts and are often considered to be good targets for drug development. Sulfur-containing-amino-acid metabolism represents one such divergent metabolic pathway, namely, the cysteine biosynthetic pathway and methionine gamma-lyase-mediated catabolism of sulfur-containing amino acids, which are present in T. vaginalis and E. histolytica but absent in G. intestinalis. These pathways are potentially exploitable for development of drugs against amoebiasis and trichomoniasis. For instance, L-trifluoromethionine, which is catalyzed by methionine gamma-lyase and produces a toxic product, is effective against T. vaginalis and E. histolytica parasites in vitro and in vivo and may represent a good lead compound. In this review, we summarize the biology of these microaerophilic parasites, their clinical manifestation and epidemiology of disease, chemotherapeutics, the modes of action of representative drugs, and problems related to these drugs, including drug resistance. We further discuss our approach to exploit unique sulfur-containing-amino-acid metabolism, focusing on development of drugs against E. histolytica.
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Affiliation(s)
- Vahab Ali
- Department of Parasitology, Gunma University Graduate School of Medicine, 3-39-22 Showa-machi, Maebashi, Gunma 371-8511, Japan
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Westrop GD, Goodall G, Mottram JC, Coombs GH. Cysteine biosynthesis in Trichomonas vaginalis involves cysteine synthase utilizing O-phosphoserine. J Biol Chem 2006; 281:25062-75. [PMID: 16735516 PMCID: PMC2645516 DOI: 10.1074/jbc.m600688200] [Citation(s) in RCA: 58] [Impact Index Per Article: 3.1] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/06/2022] Open
Abstract
Trichomonas vaginalis is an early divergent eukaryote with many unusual biochemical features. It is an anaerobic protozoan parasite of humans that is thought to rely heavily on cysteine as a major redox buffer, because it lacks glutathione. We report here that for synthesis of cysteine from sulfide, T. vaginalis relies upon cysteine synthase. The enzyme (TvCS1) can use either O-acetylserine or O-phosphoserine as substrates. The K(m) values of the enzyme for sulfide are very low (0.02 mm), suggesting that the enzyme may be a means of ensuring that sulfide in the parasite is maintained at a low level. T. vaginalis appears to lack serine acetyltransferase, the source of O-acetylserine in many cells, but has a functional 3-phosphoglycerate dehydrogenase and an O-phosphoserine aminotransferase that together result in the production of O-phosphoserine, suggesting that this is the physiological substrate. TvCS1 can also use thiosulfate as substrate. Overall, TvCS1 has substrate specificities similar to those reported for cysteine synthases of Aeropyrum pernix and Escherichia coli, and this is reflected by sequence similarities around the active site. We suggest that these enzymes are classified together as type B cysteine synthases, and we hypothesize that the use of O-phosphoserine is a common characteristic of these cysteine synthases. The level of cysteine synthase in T. vaginalis is regulated according to need, such that parasites growing in an environment rich in cysteine have low activity, whereas exposure to propargylglycine results in elevated cysteine synthase activity. Humans lack cysteine synthase; therefore, this parasite enzyme could be an exploitable drug target.
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Affiliation(s)
- Gareth D. Westrop
- Division of Infection & Immunity, Institute of Biomedical and Life Sciences and Wellcome Centre for Molecular Parasitology, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow G12 8TA, Scotland, UK
| | - Gordon Goodall
- Division of Infection & Immunity, Institute of Biomedical and Life Sciences and Wellcome Centre for Molecular Parasitology, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow G12 8TA, Scotland, UK
| | - Jeremy C. Mottram
- Division of Infection & Immunity, Institute of Biomedical and Life Sciences and Wellcome Centre for Molecular Parasitology, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow G12 8TA, Scotland, UK
| | - Graham H. Coombs
- Division of Infection & Immunity, Institute of Biomedical and Life Sciences and Wellcome Centre for Molecular Parasitology, Glasgow Biomedical Research Centre, University of Glasgow, Glasgow G12 8TA, Scotland, UK
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Pütz S, Gelius-Dietrich G, Piotrowski M, Henze K. Rubrerythrin and peroxiredoxin: two novel putative peroxidases in the hydrogenosomes of the microaerophilic protozoon Trichomonas vaginalis. Mol Biochem Parasitol 2005; 142:212-23. [PMID: 15904985 DOI: 10.1016/j.molbiopara.2005.04.003] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/02/2004] [Revised: 03/22/2005] [Accepted: 04/11/2005] [Indexed: 10/25/2022]
Abstract
The parasitic flagellate Trichomonas vaginalis contains hydrogenosomes, anaerobic organelles related to mitochondria, that generate ATP from the fermentative conversion of pyruvate to acetate, CO2 and molecular hydrogen. Although an essentially anaerobic organism, Trichomonas encounters low oxygen concentrations in its natural habitat and has to protect itself, and especially the oxygen-sensitve enzymes of hydrogenosomal metabolism, from oxidative damage. We have identified two novel proteins in the hydrogenosomal proteome with strong similarity to two putative prokaryotic peroxidases, rubrerythrin and periplasmic thiol peroxidase. Both proteins have previously been found in many prokaryotes but were not known from eukaryotes, suggesting a significant prokaryotic component in the oxygen-detoxification system of trichomonad hydrogenosomes.
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Affiliation(s)
- Simone Pütz
- Institut für Botanik III, Heinrich Heine Universtität Düsseldorf, Universitätsstrasse 1, 40225 Düsseldorf, Germany
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Cudmore SL, Delgaty KL, Hayward-McClelland SF, Petrin DP, Garber GE. Treatment of infections caused by metronidazole-resistant Trichomonas vaginalis. Clin Microbiol Rev 2004; 17:783-93, table of contents. [PMID: 15489348 PMCID: PMC523556 DOI: 10.1128/cmr.17.4.783-793.2004] [Citation(s) in RCA: 195] [Impact Index Per Article: 9.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/20/2022] Open
Abstract
Infections with the sexually transmitted protozoan Trichomonas vaginalis are usually treated with metronidazole, a 5-nitroimidazole drug derived from the antibiotic azomycin. Metronidazole treatment is generally efficient in eliminating T. vaginalis infection and has a low risk of serious side effects. However, studies have shown that at least 5% of clinical cases of trichomoniasis are caused by parasites resistant to the drug. The lack of approved alternative therapies for T. vaginalis treatment means that higher and sometimes toxic doses of metronidazole are the only option for patients with resistant disease. Clearly, studies of the treatment and prevention of refractory trichomoniasis are essential. This review describes the mechanisms of metronidazole resistance in T. vaginalis and provides a summary of trichomonicidal and vaccine candidate drugs.
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Affiliation(s)
- Sarah L Cudmore
- Division of Biochemistry, Microbiology, and Immunology, Faculty of Mediine, University of Ottawa, Ontario, Canada
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Coombs GH, Westrop GD, Suchan P, Puzova G, Hirt RP, Embley TM, Mottram JC, Müller S. The amitochondriate eukaryote Trichomonas vaginalis contains a divergent thioredoxin-linked peroxiredoxin antioxidant system. J Biol Chem 2003; 279:5249-56. [PMID: 14630923 DOI: 10.1074/jbc.m304359200] [Citation(s) in RCA: 60] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023] Open
Abstract
Trichomonas is an amitochondriate parasitic protozoon specialized for an anaerobic lifestyle. Nevertheless, it is exposed to oxygen and is able to cope with the resultant oxidative stress. In the absence of glutathione, cysteine has been thought to be the major antioxidant. We now report that the parasite contains thioredoxin reductase, which functions together with thioredoxin and thioredoxin peroxidase to detoxify potentially damaging oxidants. Thioredoxin reductase and thioredoxin also reduce cystine and so may play a role in maintaining the cellular cysteine levels. The importance of the thioredoxin system as one of the major antioxidant defense mechanisms in Trichomonas was confirmed by showing that the parasite responds to environmental changes resulting in increased oxidative stress by up-regulating thioredoxin and thioredoxin peroxidases levels. Sequence data indicate that the thioredoxin reductase of Trichomonas differs fundamentally in structure from that of its human host and thus may represent a useful drug target. The protein is generally similar to thioredoxin reductases present in other lower eukaryotes, all of which probably originated through horizontal gene transfer from a prokaryote. The phylogenetic signal in thioredoxin peroxidase is weak, but evidence from trees suggests that this gene has been subject to repeated horizontal gene transfers from different prokaryotes to different eukaryotes. The data are thus consistent with the complexity hypothesis that predicts that the evolution of simple pathways such as the thioredoxin cascade are likely to be affected by horizontal gene transfer between species.
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Affiliation(s)
- Graham H Coombs
- Division of Infection & Immunity, Institute of Biomedical and Life Sciences, University of Glasgow, Joseph Black Bldg., Glasgow G12 8QQ, United Kingdom.
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Müller S, Liebau E, Walter RD, Krauth-Siegel RL. Thiol-based redox metabolism of protozoan parasites. Trends Parasitol 2003; 19:320-8. [PMID: 12855383 DOI: 10.1016/s1471-4922(03)00141-7] [Citation(s) in RCA: 170] [Impact Index Per Article: 7.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/25/2022]
Abstract
The review considers redox enzymes of Plasmodium spp., Trypanosomatida, Trichomonas, Entamoeba and Giardia, with special emphasis on their potential use as targets for drug development. Thiol-based redox systems play pivotal roles in the success and survival of these parasitic protozoa. The synthesis of cysteine, the key molecule of any thiol metabolism, has been elucidated in trypanosomatids and anaerobes. In trypanosomatids, trypanothione replaces the more common glutathione system. The enzymes of trypanothione synthesis have recently been identified. The role of trypanothione in the detoxification of reactive oxygen species is reflected in the multiplicity of trypanothione-dependent peroxidases. In Plasmodium falciparum, the crystal structures of glutathione reductase and glutamate dehydrogenase are now available; another drug target, thioredoxin reductase, has been demonstrated to be essential for the malarial parasite.
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Affiliation(s)
- Sylke Müller
- Division of Biological Chemistry and Molecular Microbiology, School of Life Sciences, University of Dundee, DD1 5EH, Dundee, UK
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Hayashi H, Sakai H, Minakuchi-Fujiwara W, Takayama M, Nakamura-Murata M, Kamo R, Funakoshi K, Fukumoto K, Kanemaru K, Nakagawa H, Oyama Y, Shinohara N, Ito Y. Cytokinesis arrest and nuclear fission in low density populations of trichomonad protozoan. Zoolog Sci 2002; 19:1089-94. [PMID: 12426470 DOI: 10.2108/zsj.19.1089] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
Cell growth of anaerobic protozoan Tritrichomonas foetus was analyzed. This protozoan usually proliferates in extremely high density, but protozoan parasites were dispersed uniformly in F-bouillon medium and cell division stopped temporarily. However, nuclear fission continued and giant polynucleated cells formed. Later, cell division resumed and cells returned to normal form. In conditioned medium, cytokinesis of the dispersed parasites did not stop. Results indicated that T. foetus cells secreted an extracellular factor that influenced cytokinesis.
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Affiliation(s)
- Hiromi Hayashi
- Division of Life Science, Department of Mathematical and Natural Sciences, Faculty of Integrated Arts and Sciences, The University of Tokushima, Tokushima, Japan.
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