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

Mantilla, Brian S.; Paes, Lisvane S.; Pral, Elizabeth M.F.; Martil, Daiana E.; Thiemann, Otavio H.; Fernández-Silva, Patricio; Bastos, Erick L.; Silber, Ariel M.

Abstract

Proline is crucial for energizing critical events throughout the life cycle of Trypanosoma cruzi, the etiological agent of Chagas disease. The proline breakdown pathway consists of two oxidation steps, both of which producereducing equivalents as follows: the conversion of proline to Δ1-pyrroline-5-carboxylate (P5C), and the subsequent conversion of P5C to glutamate. We have identified and characterized the Δ1-pyrroline-5-carboxylate dehydrogenase from T. cruzi (TcP5CDH) and report here on how this enzyme contributes to a central metabolic pathway in this parasite. Size-exclusionchromatography, two-dimensional gel electrophoresis, and small angle x-ray scattering analysis of TcP5CDH revealed an oligomericstate composed of two subunits of six protomers. TcP5CDH was found to complement a yeast strain deficient in PUT2 activity,confirming the enzyme's functional role; and the biochemical parameters (Km, kcat, and kcat/Km) of the recombinant TcP5CDH were determined, exhibiting values comparable with those from T. cruzi lysates. In addition, TcP5CDH exhibited mitochondrial staining during the main stages of the T. cruzi life cycle. mRNA and enzymatic activity levels indicated the up-regulation (6-fold change) of TcP5CDH during the infectivestages of the parasite. The participation of P5C as an energy source was also demonstrated. Overall, we propose that thisenzymatic step is crucial for the viability of both replicative and infective forms of T. cruzi.

Citation

Mantilla, B. S., Paes, L. S., Pral, E. M., Martil, D. E., Thiemann, O. H., Fernández-Silva, P., Bastos, E. L., & Silber, A. M. (2015). Role of Δ1-Pyrroline-5-Carboxylate Dehydrogenase Supports Mitochondrial Metabolism and Host-Cell Invasion ofTrypanosoma cruzi. Journal of Biological Chemistry, 290(12), 7767-7790. https://doi.org/10.1074/jbc.m114.574525

• Journal Article Type: Article
• Online Publication Date: Jan 26, 2015
• Publication Date: Mar 20, 2015
• Deposit Date: Sep 11, 2019
• Publicly Available Date: Sep 17, 2019
• Journal: Journal of Biological Chemistry
• Print ISSN: 0021-9258
• Electronic ISSN: 1083-351X
• Publisher: American Society for Biochemistry and Molecular Biology
• Peer Reviewed: Peer Reviewed
• Volume: 290
• Issue: 12
• Pages: 7767-7790
• DOI: https://doi.org/10.1074/jbc.m114.574525
• Public URL: https://durham-repository.worktribe.com/output/1287424