title: Functional thioredoxin reductase from pathogenic and free-living Leptospira spp.
abstract: Low molecular mass thiols and antioxidant enzymes have essential functions to detoxify reactive oxygen and nitrogen species maintaining cellular redox balance. The metabolic pathways for redox homeostasis in pathogenic (Leptospira interrogans) and free-living (Leptospira biflexa) leptospires species were not functionally characterized. We performed biochemical studies on recombinantly produced proteins to in depth analyze kinetic and structural properties of thioredoxin reductase (LinTrxR) and thioredoxin (LinTrx) from L. interrogans, and two TrxRs (LbiTrxR1 and LbiTrxR2) from L. biflexa. All the TrxRs were characterized as homodimeric flavoproteins, with LinTrxR and LbiTrxR1 catalyzing the NADPH dependent reduction of LinTrx and DTNB. The thioredoxin system from L. interrogans was able to use glutathione disulfide, lipoamide disulfide, cystine and bis-γ-glutamyl cysteine and homologous peroxiredoxin as substrates. Classic TrxR activity of LinTrxR2 had not been evidenced in vitro, but recombinant Escherichia coli cells overexpressing LbiTrxR2 showed high tolerance to oxidative stress. The enzymatic systems herein characterized could play a key role for the maintenance of redox homeostasis and the function of defense mechanisms against reactive oxidant species in Leptospira spp. Our results contribute to the general knowledge about redox biochemistry in these bacteria, positioning TrxR as a critical molecular target for the development of new anti-leptospiral drugs.
authors.affiliation: Instituto de Agrobiotecnología del Litoral (CONICET-UNL), Centro Científico Tecnológico CONICET Santa Fe, Colectora Ruta Nac. N°168, km. 0, 3000 Santa Fe, Argentina; Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Ciudad Universitaria, Paraje El Pozo, 3000 Santa Fe, Argentina.
authors.forename: Natalia
authors.initials: N
authors.lastname: Sasoni
authors.affiliation: Instituto de Agrobiotecnología del Litoral (CONICET-UNL), Centro Científico Tecnológico CONICET Santa Fe, Colectora Ruta Nac. N°168, km. 0, 3000 Santa Fe, Argentina; Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Ciudad Universitaria, Paraje El Pozo, 3000 Santa Fe, Argentina.
authors.forename: Alberto A
authors.initials: AA
authors.lastname: Iglesias
authors.affiliation: Instituto de Agrobiotecnología del Litoral (CONICET-UNL), Centro Científico Tecnológico CONICET Santa Fe, Colectora Ruta Nac. N°168, km. 0, 3000 Santa Fe, Argentina; Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Ciudad Universitaria, Paraje El Pozo, 3000 Santa Fe, Argentina. Electronic address: saguerrero@ial.santafe-conicet.gov.ar.
authors.forename: Sergio A
authors.initials: SA
authors.lastname: Guerrero
authors.affiliation: Instituto de Agrobiotecnología del Litoral (CONICET-UNL), Centro Científico Tecnológico CONICET Santa Fe, Colectora Ruta Nac. N°168, km. 0, 3000 Santa Fe, Argentina; Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral, Ciudad Universitaria, Paraje El Pozo, 3000 Santa Fe, Argentina. Electronic address: darias@fbcb.unl.edu.ar.
authors.forename: Diego G
authors.initials: DG
authors.lastname: Arias
mesh_terms: D000975:Antioxidants / Q000378:metabolism; D004228:Dithionitrobenzoic Acid / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D019803:Glutathione Disulfide / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D007921:Leptospira interrogans / Q000378:metabolism / Q000472:pathogenicity; D007922:Leptospirosis / Q000378:metabolism* / Q000382:microbiology; D010084:Oxidation-Reduction; D018384:Oxidative Stress / Q000235:genetics*; D054464:Peroxiredoxins / Q000378:metabolism; D026361:Reactive Nitrogen Species / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D013438:Sulfhydryl Compounds / Q000737:chemistry; D013880:Thioredoxin-Disulfide Reductase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013879:Thioredoxins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
chemical_list: D000975:Antioxidants; D026361:Reactive Nitrogen Species; D017382:Reactive Oxygen Species; D011994:Recombinant Proteins; D013438:Sulfhydryl Compounds; D013879:Thioredoxins; D004228:Dithionitrobenzoic Acid; D054464:Peroxiredoxins; D013880:Thioredoxin-Disulfide Reductase; D019803:Glutathione Disulfide
keywords: Leptospira; Peroxiredoxin; Redox metabolism; Thioredoxin reductase