Identification and functional characterization of thioredoxin from Trypanosoma brucei brucei

Trypanosomes and Leishmania, the causative agents of several tropical disea ses, lack the glultathione/glutathione reductase system but have trypanothi one/ trypanothione reductase instead. The uniqueness of this thiol metaboli sm and the failure to detect thioredoxin reductases in these parasites have led to the suggestion that these protozoa lack a thioredoxin system. As pr esented here, this is not the case. A gene encoding thioredoxin has been cl oned from Trypanosoma brucei, the causative agent of African sleeping sickn ess. The single copy gene, which encodes a protein of 107 amino acid residu es, is expressed in all developmental stages of the parasite. The deduced p rotein sequence is 56% identical with a putative thioredoxin revealed by th e genome pro ject of Leishmania major. The relationship to other thioredoxi ns is low. T. brucei thioredoxin is unusual in having a calculated pi value of 8.5. The gene has been overexpressed in Escherichia coli. The recombina nt protein is a substrate of human thioredoxin reductase with a K-m value o f 6 mu M but is not reduced by trypanothione reductase, T. brucei thioredox in catalyzes the reduction of insulin by dithioerythritol, and functions as an electron donor for T. brucei ribonucleotide reductase. The parasite pro tein is the first classical thioredoxin of the order Kinetoplastida charact erized so far.