We seek to understand the impact that oxidative stress has upon organisms by addressing three overarching questions: How are reactive oxygen species formed in biological environments? What biomolecules do they damage? And how do cells defend themselves against them? We have employed E. coli as a model organism, in part because we have such a detailed understanding of its physiology and biochemistry, and in part because this facultative anaerobe can be genetically manipulated in the absence of oxygen. Our next set of specific aims addresses each of these three issues: 1. Are cytoplasmic enzymes shielded from H202 that is generated by periplasmic enzymes? Is intracellular formation of reactive oxygen species especially rapid inside aerated anaerobes? Are these species the primary factor in blocking their aerobic growth? Do antibiotics actually trigger H202 stress? 2. Does superoxide damage mononuclear iron enzymes ? Do oxidants inhibit ferrochelatase? What other targets of these oxidants can we discover through transcriptome analysis? 3. How completely can manganese-fed E. coli dispense with iron? How efficiently is manganese delivered to metalloenzymes? Do lactic-acid bacteria that routinely experience H202 stress employ manganese, rather than iron, in mononuclear non-redox enzymes? What about obligate aerobes, such as eukaryotes? These questions follow directly from the results of the current project. They are fundamental to our understanding of oxidative stress in contexts of medical interest, including the pathogenicity of obligate anaerobes, the effectiveness of antibiotic and antitumor treatments, and the mechanisms by which phagocytes suppress microbial infections.

Public Health Relevance

Toxic oxygen species are believed to underpin many phenomena of medical importance, including spontaneous carcinogenesis, autoimmune disorders, the action of radiation and chemotherapies, and the cellular immune response. The proposed work will identify types of cell damage that might serve as biomarkers to confirm or refute the role of oxidative stress in these processes, and it may illuminate the mechanisms bv which some cells escape toxicity.

National Institute of Health (NIH)
National Institute of General Medical Sciences (NIGMS)
Method to Extend Research in Time (MERIT) Award (R37)
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Special Emphasis Panel (NSS)
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Anderson, Vernon
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University of Illinois Urbana-Champaign
Schools of Arts and Sciences
United States
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Lu, Zheng; Sethu, Ramakrishnan; Imlay, James A (2018) Endogenous superoxide is a key effector of the oxygen sensitivity of a model obligate anaerobe. Proc Natl Acad Sci U S A 115:E3266-E3275
Li, Xin; Imlay, James A (2018) Improved measurements of scant hydrogen peroxide enable experiments that define its threshold of toxicity for Escherichia coli. Free Radic Biol Med 120:217-227
Lu, Zheng; Imlay, James A (2017) The Fumarate Reductase of Bacteroides thetaiotaomicron, unlike That of Escherichia coli, Is Configured so that It Does Not Generate Reactive Oxygen Species. MBio 8:
Khademian, Maryam; Imlay, James A (2017) Escherichia coli cytochrome c peroxidase is a respiratory oxidase that enables the use of hydrogen peroxide as a terminal electron acceptor. Proc Natl Acad Sci U S A 114:E6922-E6931
Imlay, James A (2015) Transcription Factors That Defend Bacteria Against Reactive Oxygen Species. Annu Rev Microbiol 69:93-108
Mancini, Stefano; Imlay, James A (2015) Bacterial Porphyrin Extraction and Quantification by LC/MS/MS Analysis. Bio Protoc 5:
Mancini, Stefano; Imlay, James A (2015) The induction of two biosynthetic enzymes helps Escherichia coli sustain heme synthesis and activate catalase during hydrogen peroxide stress. Mol Microbiol 96:744-63
Imlay, James A (2015) Diagnosing oxidative stress in bacteria: not as easy as you might think. Curr Opin Microbiol 24:124-31
Imlay, James A (2014) The mismetallation of enzymes during oxidative stress. J Biol Chem 289:28121-8
Sobota, Jason M; Gu, Mianzhi; Imlay, James A (2014) Intracellular hydrogen peroxide and superoxide poison 3-deoxy-D-arabinoheptulosonate 7-phosphate synthase, the first committed enzyme in the aromatic biosynthetic pathway of Escherichia coli. J Bacteriol 196:1980-91

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