Structural and functional characterization of the Pseudomonas hydroperoxide resistance protein Ohr.
Bacteria have developed complex strategies to detoxify and repair damage caused by reactive oxygen species. These compounds, produced during bacterial aerobic respiration as well as by the host immune system cells as a defense mechanism against the pathogenic microorganisms, have the ability to damage nucleic acids, proteins and phospholipid membranes. Here we describe the crystal structure of Pseudomonas aeruginosa Ohr, a member of a recently discovered family of organic hydroperoxide resistance proteins. Ohr is a tightly folded homodimer, with a novel alpha/beta fold, and contains two active sites located at the monomer interface on opposite sides of the molecule. Using in vitro assays, we demonstrate that Ohr functions directly as a hydroperoxide reductase, converting both inorganic and organic hydroperoxides to less toxic metabolites. Site-directed mutagenesis confirms that the two conserved cysteines in each active site are essential for catalytic activity. We propose that the Ohr catalytic mechanism is similar to that of the structurally unrelated peroxiredoxins, directly utilizing highly reactive cysteine thiol groups to elicit hydroperoxide reduction.
Lesniak
Jacob
J
Joan and Sanford I.Weill Graduate School of Medical Sciences of Cornell University, New York City, NY 10021, USA.
Barton
William A
WA
Nikolov
Dimitar B
DB
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry* / Q000502:physiology*; D001665:Binding Sites; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D019281:Dimerization; D004305:Dose-Response Relationship, Drug; D004351:Drug Resistance; D004926:Escherichia coli / Q000378:metabolism; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D018384:Oxidative Stress; D009195:Peroxidase / Q000378:metabolism; D010544:Peroxidases / Q000378:metabolism; D054464:Peroxiredoxins; D011487:Protein Conformation; D017510:Protein Folding; D017433:Protein Structure, Secondary; D011549:Pseudomonas / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013997:Time Factors
D001426:Bacterial Proteins; C112257:organic hydroperoxide resistance protein, Xanthomonas campestris; D006861:Hydrogen Peroxide; D010544:Peroxidases; D054464:Peroxiredoxins; D009195:Peroxidase