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A New Flavin Radical Signal in the Na+-pumping NADH:Quinone Oxidoreductase from Vibrio cholerae.
- Source :
-
Journal of Biological Chemistry . 12/1/2006, Vol. 281 Issue 48, p36482-36491. 10p. 2 Diagrams, 4 Charts, 8 Graphs. - Publication Year :
- 2006
-
Abstract
- The Na+-pumping NADH-ubiquinone oxidoreductase has six polypeptide subunits (NqrA-F) and a number of redox cofactors, including a noncovalently bound FAD and a 2Fe-2S center in subunit F, covalently bound FMNs in subunits B and C, and a noncovalently bound riboflavin in an undisclosed location. The FMN cofactors in subunits B and C are bound to threonine residues by phosphoester linkages. A neutral flavin-semiquinone radical is observed in the oxidized enzyme, whereas an anionic flavin-semiquinone has been reported in the reduced enzyme. For this work, we have altered the binding ligands of the FMNs in subunits B and C by replacing the threonine ligands with other amino acids, and we studied the resulting mutants by EPR and electron nuclear double resonance spectroscopy. We conclude that the sodium-translocating NADH:quinone oxidoreductase forms three spectroscopically distinct flavin radicals as follows: 1) a neutral radical in the oxidized enzyme, which is observed in all of the mutants and most likely arises from the riboflavin; 2) an anionic radical observed in the fully reduced enzyme, which is present in wild type, and the NqrC-T225Y mutant but not the NqrB-T236Y mutant; 3) a second anionic radical, seen primarily under weakly reducing conditions, which is present in wild type, and the NqrB-T236Y mutant but not the NqrC-T225Y mutant. Thus, we can tentatively assign the first anionic radical to the FMN in subunit B and the second to the FMN in subunit C. The second anionic radical has not been reported previously. In electron nuclear double resonance spectra, it exhibits a larger line width and larger 8α-methyl proton splittings, compared with the first anionic radical. [ABSTRACT FROM AUTHOR]
- Subjects :
- *FLAVINS
*COENZYMES
*PIGMENTS
*QUINONE
*OXIDOREDUCTASES
*ENZYMES
Subjects
Details
- Language :
- English
- ISSN :
- 00219258
- Volume :
- 281
- Issue :
- 48
- Database :
- Academic Search Index
- Journal :
- Journal of Biological Chemistry
- Publication Type :
- Academic Journal
- Accession number :
- 23576090
- Full Text :
- https://doi.org/10.1074/jbc.M605765200