The Physiology of Oxidative Stress in Bacteria

Contemporary organisms inherited their biochemistry and basic metabolic pathways from ancestors that lived in an anoxic, iron-rich world. The subsequent aeration of the planet led to problems, because oxygen is a reactive chemical that can disrupt iron-dependent enzymes. It has been the goal of our lab to detail the threats that oxygen poses, the defenses that organisms have evolved, and the situations in which oxidative stress exerts a strong impact upon microbial fate. We learned that molecular oxygen can intercept electrons as they move through redox enzymes. A mixture of superoxide and hydrogen peroxide is formed. These reactive oxygen species (ROS) are stronger oxidants than is molecular oxygen itself, and they can inappropriately disable the iron cofactors of certain enzymes. To defend themselves, cells evolved layers of defenses: scavenging enzymes that keep ROS scarce, and adaptive systems that repair damaged enzymes. ROS also threaten DNA. Hydrogen peroxide oxidizes loose iron inside the cell, forming hydroxyl radicals that can abstract electrons from DNA. Cells protect themselves by limiting the loose-iron pool and by maintaining a fleet of DNA-repair enzymes. This view is coherent and explains a lot of phenomena, but important issues are unsettled. First, we showed that different organisms produce ROS at different rates and that this variability influences oxygen sensitivity. But we do not know why ROS production varies. We are working to identify the particular enzymes that are predisposed to leak electrons to oxygen; we anticipate that ROS production will be highest in organisms that in which such enzymes are abundant. Second, we want to know which DNA repair enzymes act upon oxidative lesions. Until now, many repair mutants have not shown ROS sensitivity—but we recently developed a better understanding of how to obtain a relevant phenotype. This approach has already yielded surprising insights and suggests that oxygen was an impetus to the evolution of familiar DNA defenses. The most obvious example of oxidative stress is the phenomenon of obligate anaerobiosis. We found that aeration generates overwhelming ROS in a model anaerobe, poisoning the vulnerable enzyme families that had been identified previously. However, molecular oxygen itself also directly attacks several special enzymes that are critical for anaerobic fitness. As a test of our understanding, we aim to recapitulate evolution and fix these trouble points one by one, by engineering changes that nudge an anaerobic bacterium toward oxygen tolerance. Finally, we detailed how one redox-active antibiotic oxidizes DNA via mechanism that is shielded from cellular defenses. We will test whether other clinical antibiotics/antitumor agents do so as well. In toto, we feel we are arriving at a view of oxygen toxicity and resistance that is thorough and detailed. With that understanding may come the ability to manipulate oxidative stress in ways that are beneficial.