Thioredoxin-linked metabolism in Entamoeba histolytica.
Entamoeba histolytica, an intestinal protozoan that is the causative agent of amoebiasis, is exposed to elevated amounts of highly toxic reactive oxygen species during tissue invasion. In this work, we report the molecular cloning, from E. histolytica genomic DNA, of the genes ehtrxr and ehtrx41, respectively coding for thioredoxin reductase (EhTRXR) and thioredoxin (EhTRX41). The genes were expressed in Escherichia coli cells, and the corresponding recombinant proteins were purified and characterized. EhTRXR catalyzed the NADPH (Km=4.5 microM)-dependent reduction of 5,5'-dithiobis-(2-nitrobenzoic) acid (Km=1.7 mM), EhTRX41 (Km=3.6 microM), and E. coli TRX (Km=4.6 microM). EhTRXR and EhTRX41 could be assayed as a functional redox pair that, together with peroxiredoxin, mediate the NADPH-dependent reduction of hydrogen peroxide and tert-butyl hydroperoxide. It is proposed that this detoxifying system could be operative in vivo. Results add value to the genome project information and advise reconsideration of key metabolic pathways operating in E. histolytica.
Arias
Diego G
DG
Laboratorio de Bioquímica Microbiana, Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Ciudad Universitaria-Paraje El Pozo, 3000 Santa Fe, Argentina.
Gutierrez
César E
CE
Iglesias
Alberto A
AA
Guerrero
Sergio A
SA
D000595:Amino Acid Sequence; D000818:Animals; D003001:Cloning, Molecular; D004748:Entamoeba histolytica / Q000235:genetics / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D017125:Genes, Protozoan; D018503:Genome, Protozoan; D006861:Hydrogen Peroxide / Q000378:metabolism; D008969:Molecular Sequence Data; D009249:NADP / Q000378:metabolism; D010084:Oxidation-Reduction; D015800:Protozoan Proteins / Q000235:genetics / Q000378:metabolism*; D016415:Sequence Alignment; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D015800:Protozoan Proteins; D013879:Thioredoxins; D009249:NADP; D006861:Hydrogen Peroxide; D013880:Thioredoxin-Disulfide Reductase