Photoinitiated Electron Transfer within the Paracoccus denitrificans Cytochrome bc_1 Complex: Mobility of the Iron-Sulfur Protein Is Modulated by the Occupant of the Q_o Site

摘要

Domain rotation of the Rieske iron-sulfur protein (ISP) between the cytochrome (cyt) b and cyt c_1 redox centers plays a key role in the mechanism of the cyt bc1 complex. Electron transfer within the cyt bc_1 complex of Paracoccus denitrif icans was studied using a ruthenium dimer to rapidly photo-oxidize cyt c_1 within 1 μs and initiate the reaction. In the absence of any added quinol or inhibitor of the bc1 complex at pH 8.0, electron transfer from reduced ISP to cyt c_1 was biphasic with rate constants of k_(1f) = 6300 ± 3000 s~(-1) and k_(1s) = 640 ± 300 s~(-1) and amplitudes of 10 ± 3% and 16 ± 4% of the total amount of cyt c_1 photooxidized. Upon addition of any of the P_m type inhibitors MOA-stilbene, myxothiazol, or azoxystrobin to cyt bc1 in the absence of quinol, the total amplitude increased 2-fold, consistent with a decrease in redox potential of the ISP. In addition, the relative amplitude of the fast phase increased significantly, consistent with a change in the dynamics of the ISP domain rotation. In contrast, addition of the P_f type inhibitors JG-144 and famoxadone decreased the rate constant k_(1f) by 5-10-fold and increased the amplitude over 2-fold. Addition of quinol substrate in the absence of inhibitors led to a 2-fold increase in the amplitude of the k_(1f) phase. The effect of QH_2 on the kinetics of electron transfer from reduced ISP to cyt c_1 was thus similar to that of the P_m inhibitors and very different from that of the P_f inhibitors. The current results indicate that the species occupying the Q_o site has a significant conformational influence on the dynamics of the ISP domain rotation.