TY - JOUR
T1 - The fifth electron in the fully reduced caa3 from Thermus thermophilus is competent in proton pumping
AU - Siletsky, Sergey A.
AU - Belevich, Ilya
AU - Soulimane, Tewfik
AU - Verkhovsky, Michael I.
AU - Wikström, Mårten
N1 - Copyright © 2012 Elsevier B.V. All rights reserved.
PY - 2013/1
Y1 - 2013/1
N2 - 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 P R 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.
AB - 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 P R 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.
KW - Catalytic cycle intermediates
KW - Charge transfer steps
KW - Cytochrome c oxidase
KW - Membrane potential
KW - Thermus thermophilus
UR - http://www.scopus.com/inward/record.url?scp=84867598106&partnerID=8YFLogxK
U2 - 10.1016/j.bbabio.2012.09.013
DO - 10.1016/j.bbabio.2012.09.013
M3 - Article
C2 - 23025918
AN - SCOPUS:84867598106
SN - 0005-2728
VL - 1827
SP - 1
EP - 9
JO - Biochimica et Biophysica Acta - Bioenergetics
JF - Biochimica et Biophysica Acta - Bioenergetics
IS - 1
ER -