Structural and Biochemical Characterization of Peroxiredoxin Q beta from Xylella fastidiosa CATALYTIC MECHANISM AND HIGH REACTIVITY
Contribuinte(s) |
UNIVERSIDADE DE SÃO PAULO |
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Data(s) |
20/10/2012
20/10/2012
2010
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Resumo |
The phytopathogenic bacterium Xylella fastidiosa is the etiological agent of various plant diseases. To survive under oxidative stress imposed by the host, microorganisms express antioxidant proteins, including cysteine-based peroxidases named peroxiredoxins. This work is a comprehensive analysis of the catalysis performed by PrxQ from X. fastidiosa (XfPrxQ) that belongs to a peroxiredoxin class still poorly characterized and previously considered as moderately reactive toward hydroperoxides. Contrary to these assumptions, our competitive kinetics studies have shown that the second-order rate constants of the peroxidase reactions of XfPrxQ with hydrogen peroxide and peroxynitrite are in the order of 107 and 106 M(-1) s(-1), respectively, which are as fast as the most efficient peroxidases. The XfPrxQ disulfides were only slightly reducible by dithiothreitol; therefore, the identification of a thioredoxin system as the probable biological reductant of XfPrxQ was a relevant finding. We also showed by site-specific mutagenesis and mass spectrometry that an intramolecular disulfide bond between Cys-47 and Cys-83 is generated during the catalytic cycle. Furthermore, we elucidated the crystal structure of XfPrxQ C47S in which Ser-47 and Cys-83 lie similar to 12.3 angstrom apart. Therefore, significant conformational changes are required for disulfide bond formation. In fact, circular dichroism data indicated that there was a significant redox-dependent unfolding of alpha-helices, which is probably triggered by the peroxidatic cysteine oxidation. Finally, we proposed a model that takes data from this work as well data as from the literature into account. Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP) FAPESP Fundacao de Amparo a Pesquisa do Estado de Sao Paulo CNPq Conselho Nacional de Desenvolvimento Cientifico e Tecnologico Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq) Brazilian Synchrotron Light Laboratory (LNLS)[D03B-MX1 7568] Brazilian Synchrotron Light Laboratory (LNLS) |
Identificador |
JOURNAL OF BIOLOGICAL CHEMISTRY, v.285, n.21, p.16051-16065, 2010 0021-9258 http://producao.usp.br/handle/BDPI/27782 10.1074/jbc.M109.094839 |
Idioma(s) |
eng |
Publicador |
AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC |
Relação |
Journal of Biological Chemistry |
Direitos |
restrictedAccess Copyright AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC |
Palavras-Chave | #TRYPAREDOXIN PEROXIDASE #GLUTATHIONE PEROXIDASES #THIOREDOXIN PEROXIDASE #GENE FAMILY #PROTEIN BCP #CYSTEINE #PEROXYNITRITE #ARABIDOPSIS #EXPRESSION #OXIDATION #Biochemistry & Molecular Biology |
Tipo |
article original article publishedVersion |