Trypanosoma brucei: The effect of glycerol on the anaerobic metabolism of glucose

Hammond, David J.; Bowman, I.B.R.

doi:10.1016/0166-6851(80)90032-8

Cited by 36 publications

(16 citation statements)

References 14 publications

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“…Initially identified in intact or permeabilized cells (21)(22)(23), these organelles have been isolated (17)(18)(19)24) and found to have a high density and a high content of phosphorus, calcium, magnesium, sodium, and zinc (18,19,24). Our previous findings of large amounts of inorganic pyrophosphate in T. cruzi (10) are extended here to report the essentially complete 31 P NMR spectroscopic assignments of extracts of T. cruzi epimastigotes and their acidocalcisomes, together with the first reports of the 31 P NMR spectra and detailed assignments of whole cell and acidocalcisomal perchloric acid extracts of T. brucei and Leishmania major.…”

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confidence: 50%

“…The two most prominent resonances in the bloodstream form spectrum (A and E) occur at 4.36 and 0.03 ppm and are assigned to snglycerol-3-phosphate (G3P) 1 and glycerol-3-phosphoryl choline (GPC) based on their chemical shifts (29) and the results of co-addition of authentic G3P and GPC (data not shown). The presence of large amounts of G3P can be attributed to the well known specialized metabolism of the long slender form under anaerobic conditions (31). During centrifugation of the parasites, a decrease in the oxygen tension could lead to inactivation of the terminal mitochondrial oxidase (32) and the accumulation of G3P.…”

Section: Resultsmentioning

confidence: 99%

“…To test this idea in more detail, acidocalcisomes were isolated from T. brucei, T. cruzi, and L. major using an iodixanol (OptiPrep) density gradient (28). The densest fraction, corresponding to the acidocalcisomes, was then extracted with perchloric acid, treated with 20 mM EDTA to chelate the divalent metal cations, and observed by high resolution 31 P NMR (Fig. 3, A-D).…”

Section: Resultsmentioning

confidence: 99%

“…The specific assignments of individual resonances were initially based on published chemical shifts (29) and 1 H-31 P and 31 P-31 P scalar couplings, and were then confirmed by co-addition of authentic reference compounds. The chemical shifts of all 31 P spectra were referenced to an 85% phosphoric acid external reference at 0 ppm (10) The insets show expansions of the upfield (Ϫ4 to Ϫ23 ppm) region of each spectrum, and essentially complete resonance assignments are given in Table I. the FELIX 98 software package (Molecular Simulations Inc., San Diego, CA) running on a Silicon Graphics (SGI, Mountain View, CA) O2 workstation, and typically included base-line correction and 3-Hz exponential line broadening prior to Fourier transformation. Overall condensed inorganic phosphate levels were estimated from the NMR spectra by comparison with the ␣-ATP resonance (peak L, Ϫ10.8 ppm).…”

Section: Methodsmentioning

confidence: 99%

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31P NMR Spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major

Moreno

Urbina²,

Oldfield³

et al. 2000

Journal of Biological Chemistry

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High resolution31 P nuclear magnetic resonance spectra at 303.6 MHz (corresponding to a 1 H resonance frequency of 750 MHz) have been obtained of perchloric acid extracts of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major, the causative agents of African sleeping sickness, Chagas' disease, and leishmaniasis. Essentially complete assignments have been made based on chemical shifts and by direct addition of authentic reference compounds. The results indicate the presence of high levels of short chain condensed polyphosphates: di-, tri-, tetra-, and pentapolyphosphate.31 P NMR spectra of purified T. brucei, T. cruzi, and L. major acidocalcisomes, calcium and phosphorus storage organelles, indicate that polyphosphates are abundant in these organelles and have an average chain length of 3.11-3.39 phosphates. In the context of the recent discovery of several pyrophosphate-utilizing enzymes in trypanosomatids, the presence of these inorganic polyphosphates implies a critical role for these molecules in these parasites and a potential new route to chemotherapy.Infections caused by trypanosomatid parasites continue to be among the most widespread diseases in developing nations, and the increasing fraction of immunocompromised individuals in the global population further contributes to morbidity and economic loss. New chemotherapeutic leads based on unique aspects of parasite biology are therefore of great interest, particularly in the case of emerging resistance to traditional treatments.One approach to the discovery of new drug targets is to identify metabolic pathways in parasites that are essential for parasite survival but do not have an equivalent in the host. Then, one can pursue specific inhibitors of such metabolic activities as possible means of controlling the parasites without damaging the host. NMR spectroscopy has already been used by several groups to investigate the carbohydrate and/or phosphate metabolism of protozoa, including Crithidia fasciculata (1), Entamoeba histolytica (2), Trypanosoma cruzi (3, 4), Trypanosoma brucei (5, 6), Leishmania pifanoi (7), Babesia microti (8), and the free-living ciliate Tetrahymena pyriformis (9). In Tetrahymena, large levels of pyrophosphate were found, and more recently we have found similarly large levels of pyrophosphate in T. cruzi (10). These observations are of interest since a number of protozoa are already known to utilize pyrophosphate in addition to ATP in their high energy phosphate metabolic processes. For example, E. histolytica possesses a pyrophosphatedependent phosphofructokinase (11), a pyruvate-phosphate dikinase (12), a phosphoenolpyruvate carboxytransphosphorylase (13), and a pyrophosphate-acetate kinase (14). Several of these pyrophosphate-utilizing enzymes have also been discovered in other parasitic protists (see, e.g., the discussion in Ref. 15), and more recently a pyruvate-phosphate dikinase (16) and a plantlike vacuolar proton-translocating pyrophosphatase (17-19) were discovered in many trypanosomatids as well.An unusual character...

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confidence: 50%

Section: Resultsmentioning

confidence: 99%

Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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31P NMR Spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major

Moreno

Urbina²,

Oldfield³

et al. 2000

Journal of Biological Chemistry

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“…Since there is neither a cytosolic glycerol kinase (GK) nor any significant activity of glycerol 3-phosphatase (5, 7), all Gly-3-P is shuttled back via DHAP to the mitochondria and converted to pyruvate, leading to the production of two molecules of pyruvate per molecule of glucose consumed. Under anaerobic conditions, equimolar amounts of pyruvate and glycerol are produced due to requirements of glycosomal redox and ATP balance (1,5,(7)(8)(9)(10). This decreases the ATP production to one molecule of ATP per glucose.…”

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confidence: 99%

What Controls Glycolysis in Bloodstream Form Trypanosoma brucei?

Bakker¹,

Michels²,

Opperdoes³

et al. 1999

Journal of Biological Chemistry

169

158

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On the basis of the experimentally determined kinetic properties of the trypanosomal enzymes, the question is addressed of which step limits the glycolytic flux in bloodstream form Trypanosoma brucei. There appeared to be no single answer; in the physiological range, control shifted between the glucose transporter on the one hand and aldolase (ALD), glyceraldehyde-3-phosphate dehydrogenase (GAPDH), phosphoglycerate kinase (PGK), and glycerol-3-phosphate dehydrogenase (GDH) on the other hand. The other kinases, which are often thought to control glycolysis, exerted little control; so did the utilization of ATP.We identified potential targets for anti-trypanosomal drugs by calculating which steps need the least inhibition to achieve a certain inhibition of the glycolytic flux in these parasites. The glucose transporter appeared to be the most promising target, followed by ALD, GDH, GAPDH, and PGK. By contrast, in erythrocytes more than 95% deficiencies of PGK, GAPDH, or ALD did not cause any clinical symptoms (Schuster, R. and Holzhü tter, H.-G. (1995) Eur. J. Biochem. 229, 403-418). Therefore, the selectivity of drugs inhibiting these enzymes may be much higher than expected from their molecular effects alone. Quite unexpectedly, trypanosomes seem to possess a substantial overcapacity of hexokinase, phosphofructokinase, and pyruvate kinase, making these "irreversible" enzymes mediocre drug targets.

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Antiprotozoal Agents

Friès¹,

Fairlamb²

2003

Burger's Medicinal Chemistry and Drug Discovery

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The chapter provides coverage of all drugs currently used for the treatment of human African trypanosomiasis (African sleeping sickness), American trypanosomiases (Chagas' disease), and leishmaniasis. Collectively, this group of diseases is caused by flagellated protozoan of the order Kinetoplastida. The drugs currently used to treat these diseases include melarsoprol, suramin, pentamidine, organic antimonials, and several nitroheterocyclic agents. The agents have all been in use for 25–75 years. They have a high degree of toxicity and the first four in the preceding list must be given by parenteral injection over a 2‐ to 6‐week time period. Tolerance has developed to most of the drugs in at least some geographical areas. Eflornithine is the only agent approved in the last 50 years for the treatment of African trypanosomiasis, although it is expensive, requires prolonged systemic dosage, and is effective only against T. brucei gambiense . Megazol, trybazine, DB289, and CGP 40215A have all yielded encouraging results in in vivo preclinical studies against African trypanosomiasis. All four of these agents have entered or are scheduled to enter clinical trials. Nifurtimox and benznidazole are the only agents approved to treat Chagas' disease. The agents are toxic, rarely produce long‐term cures, and resistance to them has developed. The third‐generation triazole antifungal agents posaconazole and UR‐9825 have given encouraging preclinical results against Chagas' disease. Meltefosine has been found to be orally active and to produce 98% cure rates of visceral leishmaniasis. New classes of experimental antitrypanosomal agents include the bisphosphonates, which disrupt lipid metabolism, and cysteine protease inhibitors. Agents from both of these mechanistic classes are being advanced as potential antitrypanosomal agents. Kinetoplastid biochemistry offers numerous opportunities for the development of chemotherapeutic agents having a high degree of selective toxicity. Trypanothione, the N 1 , N 8 ‐bis(glutathionyl)spermidine metabolite that replaces glutathione in redox protection reactions in kinetoplastids, is one hopeful target for drug design. A second possible target is the kinetoplastid organelle named the glycosome. African trypanosomes and perhaps other kinetoplastids are highly dependent on glycolysis for energy production. Enclosing the enzymes for glycolysis in glycosomes increases the efficiency of glycolysis compared to its rate in mammalian cells. Any potent inhibitor of an enzyme contained in the glycosome, or of any of the biochemical processes associated with glycosomal function, might be an effective drug against these parasites.

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Trypanosoma brucei: The effect of glycerol on the anaerobic metabolism of glucose

Cited by 36 publications

References 14 publications

31P NMR Spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major

31P NMR Spectroscopy of Trypanosoma brucei, Trypanosoma cruzi, and Leishmania major

What Controls Glycolysis in Bloodstream Form Trypanosoma brucei?

Antiprotozoal Agents

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