In Trypanosoma cruzi, the etiological agent of Chagas disease, the amino acid proline participates in processes related to T. cruzi survival and infection, such as ATP production, cell differentiation, host-cell invasion, and in protection against osmotic, nutritional, and thermal stresses and oxidative imbalance. However, little is known about proline biosynthesis in this parasite. Δ1-Pyrroline-5-carboxylate reductase (P5CR, EC 1.5.1.2) catalyzes the biosynthesis of proline from Δ1-pyrroline-5-carboxylate (P5C) with concomitant NADPH oxidation. Herein, we show that unlike other eukaryotes, T. cruzi biosynthesizes proline from P5C, which is produced exclusively from glutamate. We found that TcP5CR is an NADPH-dependent cytosolic enzyme with a for P5C of 27.7 μM and with a higher expression in the insect-resident form of the parasite. High concentrations of the co-substrate NADPH partially inhibited TcP5CR activity, prompting us to analyze multiple kinetic inhibition models. The model that best explained the obtained data included a non-competitive substrate inhibition mechanism (). Therefore, TcP5CR is a candidate as a regulatory factor of this pathway. Finally, we show that P5C can exit trypanosomatid mitochondria in conditions that do not compromise organelle integrity. These observations, together with previously reported results, lead us to propose that in T. cruzi TcP5CR participates in a redox shuttle between the mitochondria and the cytoplasm. In this model, cytoplasmic redox equivalents from NADPH pools are transferred to the mitochondria using proline as a reduced metabolite, and shuttling to fuel electrons to the respiratory chain through proline oxidation by its cognate dehydrogenase.
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May 2020
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Cover Image
The cover image shows high resolution of the 20 lowest energy structures of kringle 2 of human plasminogen to AGL55-NS88.2 (top) and KT155-SS1448 (bottom). To learn more about this, see the article by Qiu and colleagues (pp. 1613–1630). The image was provided by Francis J. Castellino.
Research Article|
May 21 2020
Trypanosoma cruzi synthesizes proline via a Δ1-pyrroline-5-carboxylate reductase whose activity is fine-tuned by NADPH cytosolic pools
Letícia Marchese;
Letícia Marchese
1Laboratory of Biochemistry of Tryps - LaBTryps, Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
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Karel Olavarria;
Karel Olavarria
2Department of Microbiology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
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Brian Suarez Mantilla;
Brian Suarez Mantilla
*
1Laboratory of Biochemistry of Tryps - LaBTryps, Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
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Carla Cristi Avila;
Carla Cristi Avila
3Center of Toxins, Immune Response and Cell Signaling, Butantan Institute, 05503-900 São Paulo, SP, Brazil
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Rodolpho Ornitiz Oliveira Souza;
Rodolpho Ornitiz Oliveira Souza
1Laboratory of Biochemistry of Tryps - LaBTryps, Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
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Flávia Silva Damasceno;
Flávia Silva Damasceno
1Laboratory of Biochemistry of Tryps - LaBTryps, Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
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Maria Carolina Elias;
Maria Carolina Elias
3Center of Toxins, Immune Response and Cell Signaling, Butantan Institute, 05503-900 São Paulo, SP, Brazil
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Ariel Mariano Silber
1Laboratory of Biochemistry of Tryps - LaBTryps, Department of Parasitology, Institute of Biomedical Sciences, University of São Paulo - USP, 05508-000 São Paulo, Brazil
Correspondence: Ariel Mariano Silber (asilber@usp.br)
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Biochem J (2020) 477 (10): 1827–1845.
Article history
Received:
March 23 2020
Revision Received:
April 19 2020
Accepted:
April 20 2020
Accepted Manuscript online:
April 21 2020
Citation
Letícia Marchese, Karel Olavarria, Brian Suarez Mantilla, Carla Cristi Avila, Rodolpho Ornitiz Oliveira Souza, Flávia Silva Damasceno, Maria Carolina Elias, Ariel Mariano Silber; Trypanosoma cruzi synthesizes proline via a Δ1-pyrroline-5-carboxylate reductase whose activity is fine-tuned by NADPH cytosolic pools. Biochem J 29 May 2020; 477 (10): 1827–1845. doi: https://doi.org/10.1042/BCJ20200232
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