title: Metabolic control analysis of the Trypanosoma cruzi peroxide detoxification pathway identifies tryparedoxin as a suitable drug target.
abstract: BACKGROUND
  The principal oxidative-stress defense in the human parasite Trypanosoma cruzi is the tryparedoxin-dependent peroxide detoxification pathway, constituted by trypanothione reductase (TryR), tryparedoxin (TXN), tryparedoxin peroxidase (TXNPx) and tryparedoxin-dependent glutathione peroxidase A (GPxA). Here, Metabolic Control Analysis (MCA) was applied to quantitatively prioritize drug target(s) within the pathway by identifying its flux-controlling enzymes.
  
  
  METHODS
  The recombinant enzymes were kinetically characterized at physiological pH/temperature. Further, the pathway was in vitro reconstituted using enzyme activity ratios and fluxes similar to those observed in the parasites; then, enzyme and substrate titrations were performed to determine their degree of control on flux. Also, kinetic characterization of the whole pathway was performed.
  
  
  RESULTS
  Analyses of the kinetic properties indicated that TXN is the less efficient pathway enzyme derived from its high Kmapp for trypanothione and low Vmax values within the cell. MCA established that the TXN-TXNPx and TXN-GPxA redox pairs controlled by 90-100% the pathway flux, whereas 10% control was attained by TryR. The Kmapp values of the complete pathway for substrates suggested that the pathway flux was determined by the peroxide availability, whereas at high peroxide concentrations, flux may be limited by NADPH.
  
  
  CONCLUSIONS
  These quantitative kinetic and metabolic analyses pointed out to TXN as a convenient drug target due to its low catalytic efficiency, high control on the flux of peroxide detoxification and role as provider of reducing equivalents to the two main peroxidases in the parasite.
  
  
  CONCLUSIONS
  MCA studies provide rational and quantitative criteria to select enzymes for drug-target development.
authors.affiliation: Departamento de Bioquímica, Instituto Nacional de Cardiología Ignacio Chávez, México D.F. 14080, México.
authors.forename: Zabdi
authors.initials: Z
authors.lastname: González-Chávez
authors.affiliation: Departamento de Bioquímica, Instituto Nacional de Cardiología Ignacio Chávez, México D.F. 14080, México.
authors.forename: Viridiana
authors.initials: V
authors.lastname: Olin-Sandoval
authors.affiliation: Departamento de Bioquímica, Instituto Nacional de Cardiología Ignacio Chávez, México D.F. 14080, México.
authors.forename: José Salud
authors.initials: JS
authors.lastname: Rodíguez-Zavala
authors.affiliation: Departamento de Bioquímica, Instituto Nacional de Cardiología Ignacio Chávez, México D.F. 14080, México.
authors.forename: Rafael
authors.initials: R
authors.lastname: Moreno-Sánchez
authors.affiliation: Departamento de Bioquímica, Instituto Nacional de Cardiología Ignacio Chávez, México D.F. 14080, México. Electronic address: emma_saavedra2002@yahoo.com.
authors.forename: Emma
authors.initials: E
authors.lastname: Saavedra
mesh_terms: D014355:Chagas Disease / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D016503:Drug Delivery Systems; D006801:Humans; D007700:Kinetics; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010545:Peroxides / Q000737:chemistry / Q000378:metabolism*; D015800:Protozoan Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D014349:Trypanosoma cruzi / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D010545:Peroxides; D015800:Protozoan Proteins; D011994:Recombinant Proteins; C114110:tryparedoxin; D013879:Thioredoxins; D010088:Oxidoreductases
keywords: Flux control coefficient; Peroxiredoxin; Trypanosomatid; Trypanothione; Trypanothione reductase; Tryparedoxin