- Mycobacterium tuberculosis (M. tuberculosis) is the intracellular bacterium responsible for tuberculosis disease (TD). Inside the phagosomes of activated macrophages, M. tuberculosis is exposed to cytotoxic hydroperoxides such as hydrogen peroxide, fatty acid hydroperoxides and peroxynitrite. Thus, the characterization of the bacterial antioxidant systems could facilitate novel drug developments. In this work, we characterized the product of the gene Rv1608c from M. tuberculosis, which according to sequence homology had been annotated as a putative peroxiredoxin of the peroxiredoxin Q subfamily (PrxQ B from M. tuberculosis or MtPrxQ B). The protein has been reported to be essential for M. tuberculosis growth in cholesterol-rich medium. We demonstrated the M. tuberculosis thioredoxin B/C-dependent peroxidase activity of MtPrxQ B, which acted as a two-cysteine peroxiredoxin that could function, although less efficiently, using a one-cysteine mechanism. Through steady-state and competition kinetic analysis, we proved that the net forwardMycobacterium tuberculosis (M. tuberculosis) is the intracellular bacterium responsible for tuberculosis disease (TD). Inside the phagosomes of activated macrophages, M. tuberculosis is exposed to cytotoxic hydroperoxides such as hydrogen peroxide, fatty acid hydroperoxides and peroxynitrite. Thus, the characterization of the bacterial antioxidant systems could facilitate novel drug developments. In this work, we characterized the product of the gene Rv1608c from M. tuberculosis, which according to sequence homology had been annotated as a putative peroxiredoxin of the peroxiredoxin Q subfamily (PrxQ B from M. tuberculosis or MtPrxQ B). The protein has been reported to be essential for M. tuberculosis growth in cholesterol-rich medium. We demonstrated the M. tuberculosis thioredoxin B/C-dependent peroxidase activity of MtPrxQ B, which acted as a two-cysteine peroxiredoxin that could function, although less efficiently, using a one-cysteine mechanism. Through steady-state and competition kinetic analysis, we proved that the net forward rate constant of MtPrxQ B reaction was 3 orders of magnitude faster for fatty acid hydroperoxides than for hydrogen peroxide (3x10(6) vs 6x10(3) M-1 s(-1), respectively), while the rate constant of peroxynitrite reduction was (0.6-1.4) x10(6) M-1 s(-1) at pH 7.4. The enzyme lacked activity towards cholesterol hydroperoxides solubilized in sodium deoxycholate. Both thioredoxin B and C rapidly reduced the oxidized form of MtPrxQ B, with rates constants of 0.5x10(6) and 1x10(6) M-1 s(-1), respectively. Our data indicated that MtPrxQ B is monomeric in solution both under reduced and oxidized states. In spite of the similar hydrodynamic behavior the reduced and oxidized forms of the protein showed important structural differences that were reflected in the protein circular dichroism spectra.…
MetadatenAuthor details: | Anibal M. Reyes, Diego S. Vazquez, Ari Zeida, Martin HugoORCiD, M. Dolores Pineyro, Maria Ines De Armas, Dario Estrin, Rafael Radi, Javier Santos, Madia Trujillo |
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DOI: | https://doi.org/10.1016/j.freeradbiomed.2016.10.005 |
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ISSN: | 0891-5849 |
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ISSN: | 1873-4596 |
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Pubmed ID: | https://pubmed.ncbi.nlm.nih.gov/27751911 |
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Title of parent work (English): | Free radical biology and medicine : the official journal of the Oxygen Society, a constituent member of the International Society for Free Radical Research |
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Publisher: | Elsevier |
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Place of publishing: | New York |
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Publication type: | Article |
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Language: | English |
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Year of first publication: | 2016 |
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Publication year: | 2016 |
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Release date: | 2020/03/22 |
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Tag: | Fatty acid hydroperoxides; Mycobacterium tuberculosis; Peroxidatic and resolving cysteine; Peroxiredoxin; Peroxynitrite; Thiol-dependent peroxidase; Thioredoxin |
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Volume: | 101 |
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Number of pages: | 12 |
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First page: | 249 |
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Last Page: | 260 |
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Funding institution: | Universidad de la Republica (CSIC Grupos 767); Universidad de la Republica (Espacio Interdisciplinario); CONICET [PIP-11220110100723]; ANPCyT [PICT2014-1022, PICT2013-0982]; UBACyT [20020130100097BA, 20020130100468BA]; Universidad de la Republica-CAP, Uruguay; Universidad de Buenos Aires, Argentina |
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Organizational units: | Mathematisch-Naturwissenschaftliche Fakultät / Institut für Ernährungswissenschaft |
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Peer review: | Referiert |
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