Movement of the iron-sulfur subunit beyond the ef loop of cytochrome b is required for multiple turnovers of the bc1 complex but not for single turnover Qo site catalysis.
Recent kinetics experiments using mutants of the bc(1) complex (ubihydroquinone-cytochrome c oxidoreductase) iron-sulfur subunit with modified hinge regions have revealed the crucial role played by the large scale movement of its [2Fe-2S] cluster domain during the activity of this enzyme. In particular, one of these mutants (+1Ala) with an insertion of one alanine residue in the hinge region is partially deficient in performing this movement. We found that this defect can be overcome by the appearance of a second mutation substituting the leucine at position 286 in the ef loop of cytochrome b with a phenylalanine. Detailed studies of these mutants and their derivatives revealed that the ef loop acts as a barrier that needs to be crossed for multiple turnovers of the enzyme but not for a single turnover ubihydroquinone oxidation site catalysis. These findings indicate that the movement of the iron-sulfur subunit is composed of two discrete parts: a "micro-movement" at the cytochrome b interface, during which the [2Fe-2S] cluster interacts with ubihydroquinone oxidation site occupants and catalyzes ubihydroquinone oxidation, and a "macro-movement," during which the cluster domain swings away from cytochrome b interface, crosses the ef loop, and reaches a position close to cytochrome c(1) heme, to which it ultimately transfers an electron.
Darrouzet
Elisabeth
E
Department of Biology, Plant Science Institute, University of Pennsylvania, Philadelphia, Pennsylvania 19104, USA.
Daldal
Fevzi
F
D000409:Alanine; D001665:Binding Sites; D002384:Catalysis; D003573:Cytochrome b Group / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D003574:Cytochrome c Group / Q000378:metabolism; D003624:Darkness; D014450:Electron Transport Complex III / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007506:Iron-Sulfur Proteins / Q000737:chemistry*; D007700:Kinetics; D008958:Models, Molecular; D016254:Mutagenesis, Insertional; D010084:Oxidation-Reduction; D010649:Phenylalanine; D011199:Potentiometry; D011487:Protein Conformation; D021122:Protein Subunits; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D016354:Rhodobacter capsulatus / Q000201:enzymology
D003573:Cytochrome b Group; D003574:Cytochrome c Group; D007506:Iron-Sulfur Proteins; D021122:Protein Subunits; D011994:Recombinant Proteins; D010649:Phenylalanine; D014450:Electron Transport Complex III; D000409:Alanine