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A General Method for Detecting Nitrosamide Formation in the In Vitro Metabolism of Nitrosamines by Cytochrome P450s
Published on: September 25, 2017
A genetic analysis of nitrosative stress
Matthew W Foster1, Limin Liu, Ming Zeng
1Department of Medicine, Duke University Medical Center, Durham, North Carolina 27710, USA.
Biochemistry
|January 14, 2009
Summary
Nitric oxide (NO) and S-nitrosoglutathione (GSNO) cause nitrosative stress through protein S-nitrosylation. This study reveals distinct mechanisms for NO and GSNO, highlighting the role of metalloproteins and SNO lyase activity in nitrosative stress.
Area of Science:
- Biochemistry
- Cell Biology
- Molecular Biology
Background:
- Nitrosative stress results from elevated nitric oxide (NO) and S-nitrosothiols (e.g., S-nitrosoglutathione, GSNO), impacting protein Cys thiols (S-nitrosylation) and metallocofactors.
- The precise mechanisms by which NO and GSNO induce nitrosative stress remain incompletely understood.
Purpose of the Study:
- To elucidate the distinct roles of NO and GSNO in mediating nitrosative stress.
- To investigate the intracellular equilibrium between NO and GSNO and its dependence on cellular enzymes.
- To characterize the formation and stability of protein-bound NO species under different conditions.
Main Methods:
- Utilized yeast Saccharomyces cerevisiae strains deficient in NO- and/or GSNO-consuming enzymes (flavohemoglobin and GSNO reductase).
- Quantified the effects of NO and GSNO on cell growth and protein S-nitrosylation.
- Assessed protein S-nitrosylation in the presence and absence of oxygen, and investigated the role of transition metals.
Main Results:
- Established an intracellular equilibrium between NO and GSNO, partly mediated by SNO-lyase activity.
- Demonstrated that NO induces diverse protein modifications correlated with growth inhibition, while GSNO primarily affects S-nitrosylation.
- Identified that a significant portion of NO-induced S-nitrosylation occurs independently of oxygen, relying on protein-bound transition metals.
- Found that nitrosative stress is mainly driven by S-nitrosylation of specific protein targets, including those resistant to cellular glutathione metabolism.
Conclusions:
- NO and GSNO exhibit distinct pathways in mediating nitrosative stress.
- Protein S-nitrosylation is a complex process involving multiple synthesis and degradation routes, including metalloprotein-dependent mechanisms.
- Specific, stable protein S-nitrosylation targets are key mediators of nitrosative stress.
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