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Research has been conducted on Rhodobacter sphaeroides heme-copper oxidases with the D-channel. The characterization of the oxidase's mutant has been carried out via the examination of the individual step rates in the catalytic cycle and the results indicate that the nature of the oxygenated intermediates has not been disturbed by the mutant.

Research has been conducted on the electron-transfer from heme a to heme a (sub)3. The rate constant of this transfer and the flash-induced absorbance changes in the fully reduced and mixed-valance enzymes have been investigated and are reported.

Research describes the kinetics of the oxidation of the reduced cytochrome caa(sub)3 of Rhodothermus marinus by molecular oxygen using the flow-flash technique. Results reveal the presence of an additional, fifth electron in the enzyme as shown by two additional kinetic phases.

Research has been conducted on the 'peroxy' (P) intermediate in cytochrome c oxidase. The kinetic and thermodynamic parameters for formation of the cytochrome c oxidase with dioxygen (P (sub)m) and fully reduced enzyme with oxygen (P (sub)r) in the Rhodobacter sphaeroides enzyme are discussed.

Research points out that the origin of the absorbance difference spectrum at high pH is due to the hydroxide-ligated heme a(sub)3, which represents the transition of heme a(sub)3 from a high-spin to a low-spin state. Data suggest that proton pumping may be catalyzed by the free energy of the re-reduction of heme a(sub)3 and protonation of the hydroxyl moiety.

Research discusses the formation of F intermediate at a glutamic-acid residue, (E(I-286) and its reprotonation during proton-transfer reactions in cytochrome c oxidase from Rhodobacter sphaeroides. Data for rates of internal proton transfer from (E(I-286) to the proton acceptor and the influence of pH are presented.

Investigates the ligand-binding dynamics and the reaction of cytochrome caa[sub 3] from the thermophilic bacterium Rhodothermus marinus. Characteristics of thermophilic enzymes; Observation of heme c; Comparison of the slower transmission time of bovine enzyme.

We have investigated the CO-recombination kinetics after flash photolysis of CO from the ‘half-reduced’ cytochrome c oxidase as a function of pH. In addition, the reaction was investigated in mutant enzymes in which Lys(I-362) and Ser(I-299), located approximately in the middle of the K-pathway and near the enzyme surface, respectively, were modified. Laser-flash induced dissociation of CO is followed by rapid internal electron transfer from heme a3 to a. At pH>7 this electron transfer is associated with proton release to the bulk solution (τ≅1 ms at pH 8). Thus, the CO-recombination kinetics reflects protonation events at the catalytic site. In the wild-type enzyme, below pH∼7, the main component in the CO-recombination displayed a rate of ∼20 s−1. Above pH∼7, a slow CO-recombination component developed with a rate that decreased from ∼8 s−1 at pH 8 to ∼1 s−1 at pH 10. This slow component was not observed with KM(I-362), while with the SD(I-299)/SG(I-299) mutant enzymes at each pH it was slower than with the wild-type enzyme. The results are interpreted in terms of proton release from H2O in the catalytic site after CO dissociation, followed by OH− binding to the oxidized heme a3. The CO-recombination kinetics is proposed to be determined by the protonation rate of OH− and not dissociation of OH−, i.e. the K-pathway transfers protons and not OH−. With the KM(I-362) mutant enzyme the proton is not released, i.e. OH− is not formed. With the SD(I-299)/SG(I-299) mutant enzymes the proton is released, but both the release and uptake are slowed by the mutations. During reaction of the reduced enzyme with O2, the H2O at the binuclear center is most likely involved as a proton donor in the O–O cleavage reaction. [ABSTRACT FROM AUTHOR]