Biochemical characterization of stage-specific isoforms of aspartate aminotransferases from Trypanosoma cruzi and Trypanosoma brucei

Marciano, Daniele; Llorente, Constanza; Maugeri, Dante; Fuente, Candelaria de la; Opperdoes, Frederik; Cazzulo, Juán José; Nowicki, Cristina

doi:10.1016/j.molbiopara.2008.05.005

Cited by 39 publications

(41 citation statements)

References 25 publications

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“…Interestingly, this glutamic acid is actively taken up from the extracellular medium via a single transporter (107). However, several intracellular sources of the intramitochondrial synthesis of glutamate by enzymatic action have been demonstrated (46,47,108). In addition to the oxidation of ␥GS (catalyzed by TcP5CDH), glutamate can be enzymatically generated from the transamination of L-alanine or L-aspartate, the conversion of L-histidine, or the reductive amination of ␣-ketoglutarate (␣-KG).…”

Section: Discussionmentioning

confidence: 99%

“…The enzymatic activities detected in the mitochondrial fractions were also confirmed by Western blot analysis. The mitochondrial isoform of TcASATm was previously reported as a mitochondrial matrix protein (46). Thus, the polyclonal serum anti-TcASATm (expected size 45 kDa) was reactive in T. cruzi total extracts (Te), whereas in mitochondrial (Mc) preparations, the signal was not apparent (Fig.…”

mentioning

confidence: 82%

“…Digitonin Titration-Selective digitonin permeabilization of the T. cruzi epimastigote forms was performed as described previously (46). Briefly, cell pellets (60 mg) were harvested and resuspended in 1 ml of TSB buffer (25 mM Tris-HCl, pH 7.6, 0.25 M sucrose, 1 mM EDTA, 1 mM PMSF, and 50 l of protease/ inhibitor mixture from Sigma) in the presence of increasing concentrations of digitonin (ranging from 0 to 5 mg⅐ml Ϫ1 ) (Sigma).…”

Section: Subcellular Localization Of Tcp5cdhmentioning

confidence: 99%

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Role of Δ1-Pyrroline-5-Carboxylate Dehydrogenase Supports Mitochondrial Metabolism and Host-Cell Invasion of Trypanosoma cruzi

Mantilla

Paes

Pral

et al. 2015

Journal of Biological Chemistry

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 82%

Section: Subcellular Localization Of Tcp5cdhmentioning

confidence: 99%

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Role of Δ1-Pyrroline-5-Carboxylate Dehydrogenase Supports Mitochondrial Metabolism and Host-Cell Invasion of Trypanosoma cruzi

Mantilla

Paes

Pral

et al. 2015

Journal of Biological Chemistry

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“…1G), which can be attributed to the presence of the mitochondrial form (mASAT). This activity is found in procyclic but not bloodstream forms and is highly specific for aspartate (28).…”

Section: Aromatic Ketoacids Are Produced By Transamination Catalyzed mentioning

confidence: 98%

“…Several studies indicate that T. brucei cytoplasmic aspartate aminotransferase (TbcASAT) can efficiently catalyze transamination of aromatic amino acids (27,28). Therefore, the functional role of TbcASAT in aromatic ketoacid production was investigated by conditional interference RNA (RNAi).…”

Section: Aromatic Ketoacids Are Produced By Transamination Catalyzed mentioning

confidence: 99%

Trypanosoma bruceimetabolite indolepyruvate decreases HIF-1α and glycolysis in macrophages as a mechanism of innate immune evasion

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

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The parasite Trypanasoma brucei causes African trypanosomiasis, known as sleeping sickness in humans and nagana in domestic animals. These diseases are a major burden in the 36 sub-Saharan African countries where the tsetse fly vector is endemic. Untreated trypanosomiasis is fatal and the current treatments are stagedependent and can be problematic during the meningoencephalitic stage, where no new therapies have been developed in recent years and the current drugs have a low therapeutic index. There is a need for more effective treatments and a better understanding of how these parasites evade the host immune response will help in this regard. The bloodstream form of T. brucei excretes significant amounts of aromatic ketoacids, including indolepyruvate, a transamination product of tryptophan. This study demonstrates that this process is essential in bloodstream forms, is mediated by a specialized isoform of cytoplasmic aminotransferase and, importantly, reveals an immunomodulatory role for indolepyruvate. Indolepyruvate prevents the LPS-induced glycolytic shift in macrophages. This effect is the result of an increase in the hydroxylation and degradation of the transcription factor hypoxia-inducible factor-1α (HIF-1α). The reduction in HIF-1α levels by indolepyruvate, following LPS or trypanosome activation, results in a decrease in production of the proinflammatory cytokine IL-1β. These data demonstrate an important role for indolepyruvate in immune evasion by T. brucei.immunometabolism | innate immunity | immune evasion |

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LC–MS-based absolute metabolite quantification: application to metabolic flux measurement in trypanosomes

et al. 2015

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Human African trypanosomiasis is a neglected tropical disease caused by the protozoan parasite, Trypanosoma brucei. In the mammalian bloodstream, the trypanosome’s metabolism differs significantly from that of its host. For example, the parasite relies exclusively on glycolysis for energy source. Recently, computational and mathematical models of trypanosome metabolism have been generated to assist in understanding the parasite metabolism with the aim of facilitating drug development. Optimisation of these models requires quantitative information, including metabolite concentrations and/or metabolic fluxes that have been hitherto unavailable on a large scale. Here, we have implemented an LC–MS-based method that allows large scale quantification of metabolite levels by using U-13C-labelled E. coli extracts as internal standards. Known amounts of labelled E. coli extract were added into the parasite samples, as well as calibration standards, and used to obtain calibration curves enabling us to convert intensities into concentrations. This method allowed us to reliably quantify the changes of 43 intracellular metabolites and 32 extracellular metabolites in the medium over time. Based on the absolute quantification, we were able to compute consumption and production fluxes. These quantitative data can now be used to optimise computational models of parasite metabolism.Electronic supplementary materialThe online version of this article (doi:10.1007/s11306-015-0827-2) contains supplementary material, which is available to authorized users.

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Biochemical characterization of stage-specific isoforms of aspartate aminotransferases from Trypanosoma cruzi and Trypanosoma brucei

Cited by 39 publications

References 25 publications

Role of Δ1-Pyrroline-5-Carboxylate Dehydrogenase Supports Mitochondrial Metabolism and Host-Cell Invasion of Trypanosoma cruzi

Role of Δ1-Pyrroline-5-Carboxylate Dehydrogenase Supports Mitochondrial Metabolism and Host-Cell Invasion of Trypanosoma cruzi

Trypanosoma bruceimetabolite indolepyruvate decreases HIF-1α and glycolysis in macrophages as a mechanism of innate immune evasion

LC–MS-based absolute metabolite quantification: application to metabolic flux measurement in trypanosomes

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