Cytochrome bci Inhibitors

A third, clearer explanation of the electron transfer, proton translocation cycle is given by Saratse. Each ubiquinol (QH2) molecule can donate two electrons. A hrst QH2 electron is transferred along a high-potential chain to the [2Fe-2S] center of the ISP and then to cytochrome Ci. From the cytochrome Cl site, the electron is delivered to the attached, soluble cytochrome c in the intermembrane space. A second QH2 electron is transferred to the Qi site via the cytochrome b hemes, bL and bn. This is an electrogenic step driven by the potential difference between the two b hemes. This step creates part of the proton-motive force. After two QH2 molecules are oxidized at the Qo site, two electrons have been transferred to the Qi site (where one ubiquinone (Qio) can now be reduced, requiring two protons to be translocated from the matrix space). The net effect is a translocation of two protons for each electron transferred to cytochrome c. Each explanation of the cytochrome bci Q cycle has its merits and its proponents. The reader should consult the literature for updates in this ongoing research area. 

PDB IPPJ numbering atoms involved in hydrogen bonding  

The Di Xia research group has completed X-ray crystallographic studies of cytochrome bci with a number of different inhibitors (PDB ISQB, ISQP, ISQQ, ISQV, ISQX). Crystals suitable for crystallography were obtained by co-crystallization of the inhibitor with the native enzyme. Data for two of the inhibitor-modified cytochrome bci complex (PDB ISQP and ISQX) are included in Table 7.6 and Table 7.7. Qne inhibitor-enzyme complex studied by 

TABLE 7.7 Selected Substrate/Inhibitor Contacts at Q and Qi Sites in Cytochrome bci 

PDB No. (Substrate/ Inhibitor) Ar-Ar or hydrophobic contacts hisiei (H161) (from ISP, [2Fe-2S] ligand) (A) giu271 (E271) (from cyto b, EF ioop) (A) tyr273 (Y273) (from cyto b, EF loop) (A) 

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Only a few of the many known cytochrome bci complex inhibitors will be mentioned in this chapter. Cytochrome bci inhibitors that bind at the Qo site (p side, near the intermembrane space) include myxothiazol (MYX) and stigmatellin (SMA) (see Figure 7.29). MYX and SMA inhibit electron transfer from ubiquinol (QH2) to cytochrome c (outside the bci complex) or to other domains of the bci complex. Antimycin A (see Figure 7.29), a fungicide, binds... 

In 2003, the Di Xia group published an X-ray crystallographic study of substrate and inhibitor molecules at the Qo and Qi site. They intended to compare the structure of the native enzyme (PDB INTM) with the cytochrome bci complex having substrate or inhibitors in the Qo and/or Qi sites. The complex with substrate ubiquinone UQ2 (PDB INTZ) was discussed previously in this section. The complex with the inhibitor antimycin Ai (PDB INTK) will be discussed here. A fourth complex with the ubiquinone-model... 

In 2005, the E. A. Berry group published X-ray crystallographic studies of cytochrome bci with bound inhibitors. The structure of PDB 1PP9, resolution 2.23 A, includes stigmatellin A (SMA) in the Qo site and substrate ubiquinone (UQ) in the Qi site, while PDB IPPJ, resolution 2.28 A, includes stigmatellin A in the Qo site and antimycin Ai (ANY) in the Qi site. These... 

Figure 7.30 Comparison of cytochrome bci structures (A) PDB 1PP9 with inhibitor stigmatellin and (B) PDB IPPJ with inhibitors stigmatellin and antimycin A. Visualized using The PyMOL Molecular Graphics System and ChemDraw Ultra, version 10.0. (Printed with permission of Delano Scientific, LLC and CambridgeSoft Corporation.) (See color plate)...

PDB IPPJ cytochrome bci with inhibitor stigmatellin and antibiotic antimycin.Note [2Fe-2S] centers interact with hemes in the other monomer. 

Molecular Modeling of Inhibitors at Qi and Qo Sites in Cytochrome bci Complex... 

Figure 2. Representative structures of inhibitors of the cytochrome bci complex (cf section 2). The toxophore moiety has been indicated.

Figure 8.14. Stereo view of the monomer of Complex III the cytochrome bci complex of chicken. Shown are ligands plus the Rieske Iron Protein, where the inhibitors stigmatellin and antimycin are used to show FeS center relocation on reduction. The Rieske Iron Protein anchors in one monomer and then reaches across to accept an electron at its FeS center from the Qo site of the second monomer. (A) With inhibitors, the FeS center is in position, with the tip of the globular component of the Rieske Iron Protein pointing directly downward at the level of the Qo site, where it is to be reduced by the second component of the functional homodimer. (Prepared using the crystallographic results of Zhang et al. as...

Figure 8.19. Stereo views of monomers of Complex III cytochrome bci complex of chicken with (A) and without (B) inhibitors. Protein in backbone representation with neutral residues in light gray, hydrophobics in gray, aromatics in black, and charged residues in white. (A) The tether is extended when the FeS is at Q site. (Prepared using the crystallographic results of Zhang et al. as obtained from...

Figure 2. Structures of Other Inhibitors of the Cytochrome BCi Complex.

Inhibitors binding at the Q site close to heme ba, e.g., antimycin A and hydroxy-quinoline-N-oxides. Qi site inhibitors prevent the reduction of quinone and the reoxidation of cytochrome b through the Qi site due to the coupled reaction at the Qo site, this will also inhibit the steady state reduction of cytochrome ci but not the presteady state reduction of oxidized bci complex. 

It was unexpected that the extrinsic domain of Rieske protein was found at different positions in different crystal forms. In bovine structure IQCR (see Table 2), the location of the Rieske extrinsic domain was identified to be close to cytochrome b (Xia et al., 1997). This domain was suggested to have high mobility based on further inhibitor binding experiments and anomalous scattering studies (Kim et al., 1998). In the native chicken bc structure (IBCC), the Rieske domain was found to be far away from haem Ll and close to cytochrome c. However, in the chicken bci complex co-crystallized with Qo site inhibitor stigmatellin (structure 2BCC and 3BCC) the Rieske domain was close to haem Ll and far away from cytochrome Cl, which is approximately the position found in the structure... 

Figure 18-8 Stereoscopic ribbon diagrams of the chicken bci complex (A) The native dimer. The molecular twofold axis runs vertically between the two monomers. Quinones, phospholipids, and detergent molecules are not shown for clarity. The presumed membrane bilayer is represented by a gray band. (B) Isolated close-up view of the two conformations of the Rieske protein (top and long helix at right) in contact with cytochrome b (below), with associated heme groups and bound inhibitors, stigmatellin, and antimycin. The isolated heme of cytochrome Cj (left, above) is also shown. (C) Structure of the intermembrane (external surface) domains of the chicken bc complex. This is viewed from within the membrane, with the transmembrane helices truncated at roughly the membrane surface. BaU-and-stick models represent the heme group of cytochrome Cj, the Rieske iron-sulfur cluster, and the disulfide cysteines of subunit 8. SU, subunit cyt, cytochrome. From Zhang et al ° ...

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