The inside pH determines rates of electron and proton transfer in vesicle-reconstituted cytochrome c oxidase.
Cytochrome c oxidase is the terminal enzyme in the respiratory chains of mitochondria and many bacteria where it translocates protons across a membrane thereby maintaining an electrochemical proton gradient. Results from earlier studies on detergent-solubilized cytochrome c oxidase have shown that individual reaction steps associated with proton pumping display pH-dependent kinetics. Here, we investigated the effect of pH on the kinetics of these reaction steps with membrane-reconstituted cytochrome c oxidase such that the pH was adjusted to different values on the inside and outside of the membrane. The results show that the pH on the inside of the membrane fully determines the kinetics of internal electron transfers that are linked to proton pumping. Thus, even though proton release is rate limiting for these reaction steps (Salomonsson et al., Proc. Natl. Acad. Sci. USA, 2005, 102, 17624), the transition kinetics is insensitive to the outside pH (in the range 6-9.5).
Faxén
Kristina
K
Department of Biochemistry and Biophysics, The Arrhenius Laboratories for Natural Sciences, Stockholm University, SE-106 91 Stockholm, Sweden.
Brzezinski
Peter
P
D001426:Bacterial Proteins / Q000737:chemistry / Q000378:metabolism; D003576:Electron Transport Complex IV / Q000737:chemistry / Q000378:metabolism*; D004583:Electrons; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D011487:Protein Conformation; D011522:Protons; D012242:Rhodobacter sphaeroides / Q000201:enzymology
D001426:Bacterial Proteins; D011522:Protons; D003576:Electron Transport Complex IV