Ascochlorin is a novel, specific inhibitor of the mitochondrial cytochrome bc1 complex

Abstract

Ascochlorin is an isoprenoid antibiotic that is produced by the phytopathogenic fungus Ascochyta viciae. Similar to ascofuranone, which specifically inhibits trypanosome alternative oxidase by acting at the ubiquinol binding domain, ascochlorin is also structurally related to ubiquinol. When added to the mitochondrial preparations isolated from rat liver, or the yeast Pichia (Hansenula) anomala, ascochlorin inhibited the electron transport via CoQ in a fashion comparable to antimycin A and stigmatellin, indicating that this antibiotic acted on the cytochrome bc1 complex. In contrast to ascochlorin, ascofuranone had much less inhibition on the same activities. On the one hand, like the Qi site inhibitors antimycin A and funiculosin, ascochlorin induced in H. anomala the expression of nuclear-encoded alternative oxidase gene much more strongly than the Qo site inhibitors tested. On the other hand, it suppressed the reduction of cytochrome b and the generation of superoxide anion in the presence of antimycin A3 in a fashion similar to the Qo site inhibitor myxothiazol. These results suggested that ascochlorin might act at both the Qi and the Qo sites of the fungal cytochrome bc1 complex. Indeed, the altered electron paramagnetic resonance (EPR) line shape of the Rieske iron-sulfur protein, and the light-induced time resolved cytochrome b and c reduction kinetics of Rhodobacter capsulatus cytochrome bc1 complex in the presence of ascochlorin demonstrated that this inhibitor can bind to both the Qo and Qi sites of the bacterial enzyme. Additional experiments using purified bovine cytochrome bc1 complex showed that ascochlorin inhibits reduction of cytochrome b by ubiquinone through both Qi and Qo sites. Moreover, crystal structure of chicken cytochrome bc1 complex treated with excess ascochlorin revealed clear electron densities that could be attributed to ascochlorin bound at both the Qi and Qo sites. Overall findings clearly show that ascochlorin is an unusual cytochrome bc1 inhibitor that acts at both of the active sites of this enzyme.

HDQ, a Potent Inhibitor of Plasmodium falciparum Proliferation, Binds to the Quinone Reduction Site of the Cytochrome bc 1 Complex

“…Inhibitors have been described that bind at both Q o and Q i sites. For instance, spectroscopic studies have shown that Ascochlorin acted at both sites of the bacterial and vertebrate bc 1 complex and crystallographic analyses has revealed its precise binding sites in the two quinone pockets of the chicken enzyme (3). Crystallographic analysis of the bovine bc 1 complex showed that 2-nonyl-4-hydroxyquinoline Noxide (NQNO) binds at both sites in the crystal structure (25).…”

4-O-Carboxymethyl Ascochlorin Causes ER Stress and Induced Autophagy in Human Hepatocellular Carcinoma Cells

“…Recent results show that ASC inhibits mitochondrial electron transport by binding to cytochrome bc1. The x-ray crystal structure of the complex shows ASC bound at both active sites of cytochrome bc1 (6).…”

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

“…The Q i site of cytochrome bc 1 has been far less explored in the search for antimalarial compounds and only the binding of a few compounds has been visualized directly (38)(39)(40). Several compounds have been suggested to bind the Q i site, including potential antimalarial compounds, but due to lack of structural information have not been pursued vigorously (41).…”