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Doctoral Thesis
DOI
https://doi.org/10.11606/T.42.2018.tde-02022018-151505
Document
Author
Full name
Letícia Marchese
Institute/School/College
Knowledge Area
Date of Defense
Published
São Paulo, 2017
Supervisor
Committee
Silber, Ariel Mariano (President)
Cordeiro, Artur Torres
Festuccia, William Tadeu Lara
Labriola, Leticia
Thiemann, Otavio Henrique
Title in Portuguese
Caracterização molecular e bioquímica das enzimas envolvidas na biossíntese de prolina em Trypanosoma cruzi.
Keywords in Portuguese
Trypanossoma cruzi
Biossíntese de L-prolina
Metabolismo
Transidrogenase
Tritryps
Abstract in Portuguese
Alguns organismos podem biossintetizar L-prolina (L-Pro) principalmente via L-glutamato (L-Glu). A Δ1-pirrolina-5-carboxilato (P5C) sintase (P5CS) converte o L-Glu ao intermediário da via, o P5C que uma vez formado é reduzido a L-Pro via P5C redutase (P5CR). Nesse trabalho, verificou-se a possível ocorrência dessa via em Tritryps. Em Trypanosoma cruzi toda a via é operativa, em Trypanosoma brucei está ausente, enquanto que em Leishmania amazonensis acontece parcialmente. A biossíntese de L-Pro em T. cruzi é abordada com mais detalhes. Ambas as enzimas da via foram expressas e tiveram sua localização determinada no citosol, compartimento distinto das enzimas do catabolismo de L-Pro. Ainda realizou-se a caracterização bioquímica da TcP5CR, que curiosamente sofre inibição incompetitiva pelo NADPH, e por isso é candidata a regular esse metabolismo. Por fim, evidenciou-se a saída do P5C da mitocôndria, possibilitando a existência de um mecanismo de transidrogenação via a interconversão de L-Pro em P5C e vice versa.
Title in English
Molecular and biochemical characterization of the enzymes involved in proline biosynthesis in Trypanosoma cruzi.
Keywords in English
Trypanossoma cruzi
L-Proline biosynthesis
Metabolism
Transhydrogenase
Tritryps
Abstract in English
Some organisms biosynthesize L-proline (L-Pro) mainly from L-glutamate (L-Glu). The Δ1-pyrroline-5-carboxylate (P5C) synthase (P5CS) converts L-Glu into the intermediate P5C, which in turn, is reduced to L-Pro by a P5C reductase (P5CR). In this study, we verified the possible occurrence of this pathway in Tritryps. In Trypanosoma cruzi the entire route is operative. In Trypanosoma brucei, is absent, while Leishmania amazonensis this pathway is partially operative. In T. cruzi, both enzymes were expressed and had their location determined in the cytosol, differently from those of the catabolism of L-Pro. In addition, the biochemical characterization of TcP5CR showed an uncompetitive inhibition by NADPH, and therefore it is a candidate to be a regulation point of the pathway. Finally, it was observed the exit of PC5 from the mitochondria, a condition for the existence of a transhydrogenation mechanism via the interconversion between Pro and P5C.
 
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Publishing Date
2018-02-02
 
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