Analysis of the unique trypanothione redox metabolism under pseudo-physiological conditions.

LEROUX, ALEJANDRO E.; KRAUTH-SIEGEL, R. LUISE

Sant Feliu de Guixols

Conferencia; Conference on Glutathione and Related Thiols in Living Cells; 2011

European Science Foundation - EMBO

Trypanosoma brucei is the causative agent of human African trypanosomiasis, one of the most important neglected tropical diseases. The parasite possesses a unique redox metabolism which is based on the dithiol trypanothione. This bis(glutathionyl)spermidine conjugate delivers the electrons for the synthesis of DNA precursors, the detoxification of hydroperoxides, methionine sulfoxide reduction and other thiol-dependent pathways. The first two steps of trypanothione biosynthesis are shared with the glutathione biosynthesis and are catalyzed by gamma-glutamylcysteine synthetase and glutathione synthetase. In the final step, two glutathione molecules are conjugated with one spermidine by trypanothione synthetase. The Silicon Trypanosome project aims at the creation of a comprehensive, experiment-based, multi-scale mathematical model of trypanosome physiology (Bakker et al, 2010 Parasitology). Its current objective is to expand the established glycolysis model to include the pentose phosphate pathway (main source of reducing power) and the trypanothione metabolism. As part of this collaborative project we will measure the kinetic constants of the parasite redox enzymes under pseudo-physiological conditions and determine the intracellular concentration and - where appropriate - the redox state of the proteins. For this purpose, we have defined and tested a buffer system that should mimic the cytosolic milieu of the mammalian bloodstream T. brucei as much as possible. The kinetic analysis of trypanothione synthetase, one of the key enzymes of the parasite redox metabolism, under these conditions revealed remarkable differences to the published data obtained under optimized but artificial conditions. The experimental data obtained under these pseudo-physiological conditions will form the basis for the computational analysis of the parasite biology. We expect that, in the long run, the quantitative modelling enabled by the Silicon Trypanosome will play a key role in selecting molecular targets for novel antiparasitic drugs.