Mechanism of Ubiquinol Oxidation by the bc₁ Complex:  Role of the Iron Sulfur Protein and Its Mobility

Abstract

Native structures of ubihydroquinone:cytochrome c oxidoreductase (bc 1 complex) from different sources, and structures with inhibitors in place, show a 16-22 Å displacement of the [2Fe-2S] cluster and the position of the C-terminal extrinsic domain of the iron sulfur protein. None of the structures shows a static configuration that would allow catalysis of all partial reactions of quinol oxidation. We have suggested that the different conformations reflect a movement of the subunit necessary for catalysis. The displacement from an interface with cytochrome c 1 in native crystals to an interface with cytochrome b is induced by stigmatellin or 5-n-undecyl-6-hydroxy-4,7-dioxobenzothiazole (UHDBT) and involves ligand formation between His-161 of the [2Fe-2S] binding cluster and the inhibitor. The movement is a rotational displacement, so that the same conserved docking surface on the iron sulfur protein interacts with cytochrome c 1 and with cytochrome b. The mobile extrinsic domain retains essentially the same tertiary structure, and the anchoring N-terminal tail remains in the same position. The movement occurs through an extension of a helical segment in the short linking span. We report details of the protein structure for the two main configurations in the chicken heart mitochondrial complex and discuss insights into mechanism provided by the structures and by mutant strains in which the docking at the cytochrome b interface is impaired. The movement of the iron sulfur protein represents a novel mechanism of electron transfer, in which a tethered mobile head allows electron transfer through a distance without the entropic loss from free diffusion.The ubihydroquinone:cytochrome c oxidoreductase (bc 1 complex) 1 (E.C. 1.10.2.2) and the related b 6 f complex of oxygenic photosynthesis are central components of the major electron-transfer chains that carry the energy flux of the biosphere. The bc 1 complex of the mitochondrial respiratory chain transfers electrons from ubihydroquinone (quinol) to cytochrome (cyt) c in the aqueous phase and catalyzes the coupled transfer of protons across the membrane. Our understanding of the function of these enzymes at the atomic level has been greatly aided by structures recently solved by X-ray crystallography. These include cyt f (the equivalent of cyt c 1 in the b 6 f complex) from chloroplasts (1, 2) and the Rieske iron sulfur protein (ISP) from the beef heart mitochondrial complex (3) and chloroplasts (4), all crystallized as water-soluble fragments generated by proteolysis or mutagenesis. A partial structure of the beef heart mitochondrial complex at ∼2.9 Å resolution has been published by Xia and colleagues (5, 6). However, the crystals were disordered in the region of the ISP and cytochrome c 1 , and this precluded a detailed consideration of the mechanistic role of these subunits. More complete structures for the complexes from chicken, rabbit, and beef heart mitochondria have been provided by Zhang et al. ‡ University of Illinois. § University of California. ...

Mechanism of Ubiquinol Oxidation by the bc1 Complex: Different Domains of the Quinol Binding Pocket and Their Role in the Mechanism and Binding of Inhibitors

“…(g) Meanwhile, the reduced ISP can deliver an electron to cyt c 1 by tethered diffusion (38,39) and return in the oxidized form to initiate another turnover of the Q o site.…”

“…In the presence of stigmatellin, the ISP is found closely docked at the concave interface. A strong density connection is found between the headgroup of stigmatellin and H161 of the ISP (7,11,38). We have modeled this as a H-bond between N of H161, and carbonyl and O-methyl groups of stigmatellin.…”

“…The structures have already revealed a hitherto unsuspected complexity in the reaction mechanism. The extrinsic head of the ISP serves as a mobile domain, ferrying redox equivalents between reaction interfaces on cyt c 1 and cyt b through a 22 Å movement (7,11,(38)(39)(40).…”

Exploration of Ligands to the Qi Site Semiquinone in the bc 1 Complex Using High-resolution EPR

“…The nuclear quadrupole resonance method also provides a value for the asymmetry parameter, ϭ 0.39, which allows us to calculate a value for of 1. (20), state B represents the structure of Berry and co-workers (24), and state C represents the structure determined here by ESEEM. The second electron transfer is reversible, so that formation of the semiquinone from QH 2 upon addition to the oxidized complex would be represented by reversal of transitions leading to the formation of state A from C. No attempt has been made to show the many different states of protonation of the residues involved, and the particular points of entry of H ϩ in the scheme should, therefore, be regarded as flexible and would change with pH.…”

Antimalarial 4(1H)-pyridones bind to the Q i site of cytochrome bc 1

“…There are three catalytic subunits: cytochrome b, cytochrome c 1 , and the Rieske iron-sulfur protein, which are present in all bc 1 complexes (12). The complex forms part of the respiratory chain and acts to transport H + into the intermembrane space through the oxidation and reduction of ubiquinone in the modified Q cycle (12,15,16). The Q cycle requires two distinct binding sites for the reduction and oxidation of ubiquinol and ubiquinone.…”