کد مقاله | کد نشریه | سال انتشار | مقاله انگلیسی | نسخه تمام متن |
---|---|---|---|---|
1942267 | 1052600 | 2013 | 9 صفحه PDF | دانلود رایگان |
The time-resolved kinetics of membrane potential generation coupled to oxidation of the fully reduced (five-electron) caa3 cytochrome oxidase from Thermus thermophilus by oxygen was studied in a single-turnover regime. In order to calibrate the number of charges that move across the vesicle membrane in the different reaction steps, the reverse electron transfer from heme a3 to heme a and further to the cytochrome c/CuA has been resolved upon photodissociation of CO from the mixed valence enzyme in the absence of oxygen. The reverse electron transfer from heme a3 to heme a and further to the cytochrome c/CuA pair is resolved as a single transition with τ ~ 40 μs. In the reaction of the fully reduced cytochrome caa3 with oxygen, the first electrogenic phase (τ ~ 30 μs) is linked to OO bond cleavage and generation of the PR state. The next electrogenic component (τ ~ 50 μs) is associated with the PR → F transition and together with the previous reaction step it is coupled to translocation of about two charges across the membrane. The three subsequent electrogenic phases, with time constants of ~ 0.25 ms, ~ 1.4 ms and ~ 4 ms, are linked to the conversion of the binuclear center through the F → OH → EH transitions, and result in additional transfer of four charges through the membrane dielectric. This indicates that the delivery of the fifth electron from heme c to the binuclear center is coupled to pumping of an additional proton across the membrane.
► We have resolved kinetics of membrane potential generation by caa3 cytochrome oxidase.
► Single-turnover oxidation of the fully reduced enzyme by oxygen was studied.
► The number of charges translocated coupled to the different reaction steps were identified.
► Delivery of the fifth electron to the catalytic center is coupled to proton pumping.
Journal: Biochimica et Biophysica Acta (BBA) - Bioenergetics - Volume 1827, Issue 1, January 2013, Pages 1–9