Structural Basis for the Quinone Reduction in the bc 1 Complex:  A Comparative Analysis of Crystal Structures of Mitochondrial Cytochrome bc 1 with Bound Substrate and Inhibitors at the Qi Site

Cytochrome bc 1 is an integral membrane protein complex essential to cellular respiration and photosynthesis. The Q cycle reaction mechanism of bc 1 postulates a separated quinone reduction (Qi) and quinol oxidation (Qo) site. In a complete catalytic cycle, a quinone molecule at the Qi site receives two electrons from the b H heme and two protons from the negative side of the membrane; this process is specifically inhibited by antimycin A and NQNO. The structures of bovine mitochondrial bc 1 in the presence or absence of bound substrate ubiquinone and with either the bound antimycin A1 or NQNO were determined and refined. A ubiquinone with its first two isoprenoid repeats and an antimycin A1 were identified in the Qi pocket of the substrate and inhibitor bound structures, respectively; the NQNO, on the other hand, was identified in both Qi and Qo pockets in the inhibitor complex. The two inhibitors occupied different portions of the Qi pocket and competed with substrate for binding. In the Qo pocket, the NQNO behaves similarly to stigmatellin, inducing an iron−sulfur protein conformational arrest. Extensive binding interactions and conformational adjustments of residues lining the Qi pocket provide a structural basis for the high affinity binding of antimycin A and for phenotypes of inhibitor resistance. A two-water-mediated ubiquinone protonation mechanism is proposed involving three Qi site residues His201, Lys227, and Asp228.