title: Catalytic properties, thiol pK value, and redox potential of Trypanosoma brucei tryparedoxin.
abstract: The dithiol protein tryparedoxin is a component of the unique trypanothione/trypanothione reductase metabolism of trypanosomatids and is involved in the parasite synthesis of deoxyribonucleotides and the detoxication of hydroperoxides. Tryparedoxin is a highly abundant protein in all life stages of Trypanosoma brucei, the causative agent of African sleeping sickness. As shown here, its functional properties are intermediate between those of classical thioredoxins and glutaredoxins. The redox potential of T. brucei tryparedoxin of -249 mV was determined by protein-protein redox equilibration with Escherichia coli thioredoxin. The trypanothione/tryparedoxin couple is probably the most significant factor determining the cytosolic redox potential of the parasites. The pK value of Cys(40), the first thiol in the WCPPC motif, is 7.2 as derived from the thiolate absorption at 240 nm and the rate of carboxymethylation. Alteration of the active site into that of thioredoxin (CGPC) did not affect the pK value. In contrast, in the mutant with the glutaredoxin motif (CPYC) the pK dropped to < or =4.0. The fact that the pK value of tryparedoxin coincides with the intracellular pH of the parasite may contribute to the reactivity of tryparedoxin in thiol disulfide exchange reactions.
authors.affiliation: Biochemie-Zentrum Heidelberg, Universität Heidelberg, 69120 Heidelberg, Germany.
authors.forename: Nina
authors.initials: N
authors.lastname: Reckenfelderbäumer
authors.forename: R Luise
authors.initials: RL
authors.lastname: Krauth-Siegel
mesh_terms: D000478:Alkylation; D019943:Amino Acid Substitution; D000818:Animals; D002384:Catalysis; D002851:Chromatography, High Pressure Liquid; D003683:Dehydroascorbic Acid / Q000378:metabolism; D004789:Enzyme Activation; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008291:Malate Dehydrogenase / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D010084:Oxidation-Reduction; D013054:Spectrophotometry, Atomic; D013438:Sulfhydryl Compounds / Q000378:metabolism*; D013880:Thioredoxin-Disulfide Reductase / Q000378:metabolism; D013879:Thioredoxins / Q000235:genetics / Q000378:metabolism*; D014346:Trypanosoma brucei brucei / Q000378:metabolism*
chemical_list: D013438:Sulfhydryl Compounds; C114110:tryparedoxin; D013879:Thioredoxins; D008291:Malate Dehydrogenase; D013880:Thioredoxin-Disulfide Reductase; D003683:Dehydroascorbic Acid