Role of the General Base Glu-268 in Nitroglycerin Bioactivation and Superoxide Formation by Aldehyde Dehydrogenase-2
| Autor: | Karl Gruber, Antonius C.F. Gorren, Doris Koesling, Matteo Beretta, Pravas Kumar Baral, Andreas Zeller, Michael Russwurm, Kurt Schmidt, Bernd Mayer, M. Verena Wenzl |
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| Rok vydání: | 2009 |
| Předmět: |
Receptors
Cytoplasmic and Nuclear Aldehyde dehydrogenase Dehydrogenase Nitric Oxide medicine.disease_cause Biochemistry Nitric oxide Superoxide dismutase Nitroglycerin chemistry.chemical_compound Soluble Guanylyl Cyclase Superoxides Escherichia coli medicine Animals Humans Point Mutation Lung Molecular Biology ALDH2 chemistry.chemical_classification Enzyme Catalysis and Regulation biology Superoxide Dismutase Superoxide Aldehyde Dehydrogenase Mitochondrial Cell Biology Aldehyde Dehydrogenase NAD Enzyme chemistry Guanylate Cyclase cardiovascular system biology.protein Cattle Oxidative stress circulatory and respiratory physiology |
| Zdroj: | The Journal of Biological Chemistry |
| ISSN: | 0021-9258 |
| DOI: | 10.1074/jbc.m109.005652 |
| Popis: | Mitochondrial aldehyde dehydrogenase-2 (ALDH2) plays an essential role in nitroglycerin (GTN) bioactivation, resulting in formation of NO or a related activator of soluble guanylate cyclase. ALDH2 denitrates GTN to 1,2-glyceryl dinitrate and nitrite but also catalyzes reduction of GTN to NO. To elucidate the relationship between ALDH2-catalyzed GTN bioconversion and established ALDH2 activities (dehydrogenase, esterase), we compared the function of the wild type (WT) enzyme with mutants lacking either the reactive Cys-302 (C302S) or the general base Glu-268 (E268Q). Although the C302S mutation led to90% loss of all enzyme activities, the E268Q mutant exhibited virtually unaffected rates of GTN denitration despite low dehydrogenase and esterase activities. The nucleotide co-factor NAD caused a pronounced increase in the rates of 1,2-glyceryl dinitrate formation by WT-ALDH2 but inhibited the reaction catalyzed by the E268Q mutant. GTN bioactivation measured as activation of purified soluble guanylate cyclase or release of NO in the presence of WT- or E268Q-ALDH2 was markedly potentiated by superoxide dismutase, suggesting that bioavailability of GTN-derived NO is limited by co-generation of superoxide. Formation of superoxide was confirmed by determination of hydroethidine oxidation that was inhibited by superoxide dismutase and the ALDH2 inhibitor chloral hydrate. E268Q-ALDH2 exhibited approximately 50% lower rates of superoxide formation than the WT enzyme. Our results suggest that Glu-268 is involved in the structural organization of the NAD-binding pocket but is not required for GTN denitration. ALDH2-catalyzed superoxide formation may essentially contribute to oxidative stress in GTN-exposed blood vessels. |
| Databáze: | OpenAIRE |
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