Structural and functional characterization of an organic hydroperoxide resistance protein from Mycoplasma gallisepticum.
As obligate parasites, Mycoplasma species are continuously exposed to oxidative damage due to host-generated peroxides and reactive oxygen species (ROS). In addition, the production of endogenous oxidants is believed to be a primary virulence mechanism of several Mollicute species, indicating that oxidative stress resistance is crucial to survival of these bacteria in the host milieu. Despite the abundance of oxidants at the site of infection, enzymes responsible for the detoxification of ROS have never been characterized in mycoplasmas. Here we characterize a homolog of the ohr (organic hydroperoxide resistance) family from Mycoplasma gallisepticum (encoding MGA1142). Unlike previously characterized ohr genes, the mga1142 gene is not upregulated in response to oxidative stress but displays a novel pattern of expression. Both organic and inorganic peroxides can act as substrates for MGA1142, but they are degraded with various efficiencies. Furthermore, cumene hydroperoxide, an aromatic peroxide metabolized with high efficiency by other Ohr proteins, was shown to rapidly inactivate MGA1142, accounting for the sensitivity of M. gallisepticum cells to this compound. Comparative modeling of the MGA1142 quaternary structure revealed that the active site of this molecule has a relatively wide conformation. These data indicate that the natural substrate for MGA1142 differs from that for previously characterized Ohr proteins. Triton X-114 partitioning demonstrated that MGA1142 is located in both cytosol and membrane fractions, suggesting that in vivo this molecule plays a role in the detoxification of both endogenous and exogenous peroxides. A model describing how MGA1142 is likely to be oriented in the cell membrane is presented.
Jenkins
Cheryl
C
Department of Primary Industries, Elizabeth Macarthur Agricultural Institute, Camden, NSW 2570, Australia. cheryl.jenkins@dpi.nsw.gov.au
Samudrala
Ram
R
Geary
Steven J
SJ
Djordjevic
Steven P
SP
D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015152:Blotting, Northern; D015153:Blotting, Western; D024881:Drug Resistance, Bacterial; D015180:Electrophoresis, Gel, Two-Dimensional; D015964:Gene Expression Regulation, Bacterial / Q000187:drug effects; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D008958:Models, Molecular; D008969:Molecular Sequence Data; D045686:Mycoplasma gallisepticum / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D009195:Peroxidase / Q000378:metabolism; D010802:Phylogeny; D011401:Promoter Regions, Genetic / Q000235:genetics; D017433:Protein Structure, Secondary; D017422:Sequence Analysis, DNA
D001426:Bacterial Proteins; D006861:Hydrogen Peroxide; D009195:Peroxidase