Spatial organization of redox active centers in the bovine heart ubiquinol-cytochrome c oxidoreductase.
We have examined the spatial organization of the redox active centers in the Site II segment of the bovine heart respiratory chain by using reconstituted proteoliposomes of ubiquinol-cytochrome c oxidoreductase (Complex III or cytochrome bc1 complex) and EPR techniques. 1) Mutual spin-spin interactions between intrinsic redox active centers were detected. The spin relaxation of the Rieske iron-sulfur cluster was enhanced by the paramagnetic cytochrome c1 and b566 hemes but not by cytochrome b562. 2) Relative distances of the individual redox active centers to the P-side and N-side surfaces of the reconstituted Complex III proteoliposome were measured by our paramagnetic probe method (Blum, H., Bowyer, J. R., Cusanovich, M. A., Waring, A. J., and Ohnishi, T. (1983) Biochim. Biophys. Acta 748, 418-428). The cytochrome b562 heme was shown to be close to the middle of the phospholipid bilayer, while the Rieske iron-sulfur cluster and cytochrome b566 heme were assigned to be near the P-side surface level of the membrane. This probe method is a low resolution technique from the structural viewpoint; however, it can provide direct and reliable assignment of the topographical locations of redox active centers within the membrane. This is the first direct demonstration of the transmembranous location of the two cytochrome b hemes, although electron transfer between these two hemes crosses only half of the membrane thickness. Our data support the assignment of transmembranous distribution of the redox active centers based on electrochromic measurements (Robertson, D.E., and Dutton, P.L. (1988) Biochim, Biophys. Acta 935, 273-291). The implication of these results on the mechanism of Site II energy coupling is discussed.
Ohnishi
T
T
Department of Biochemistry and Biophysics, University of Pennsylvania, Philadelphia 19104.
Schägger
H
H
Meinhardt
S W
SW
LoBrutto
R
R
Link
T A
TA
von Jagow
G
G
D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003580:Cytochromes / Q000378:metabolism; D004578:Electron Spin Resonance Spectroscopy; D014450:Electron Transport Complex III / Q000378:metabolism*; D007700:Kinetics; D008081:Liposomes; D008929:Mitochondria, Heart / Q000201:enzymology*; D008961:Models, Structural; D010084:Oxidation-Reduction; D011487:Protein Conformation; D011510:Proteolipids / Q000378:metabolism
D003580:Cytochromes; D008081:Liposomes; D011510:Proteolipids; C020485:proteoliposomes; D014450:Electron Transport Complex III