Ubiquinone (UQ) is a redox active lipid that transfers electrons from complex I or II to complex III in the electron transport chain (ETC). The long-lived Caenorhabditis elegans mutant clk-1 is unable to synthesize its native ubiquinone, and accumulates high amounts of its precursor, 5-demethoxyubiquinone-9 (DMQ(9)). In clk-1, complexes I-III activity is inhibited while complexes II-III activity is normal. We asked whether the complexes I-III defect in clk-1 was caused by: (1) a defect in the ETC; (2) an inhibitory effect of DMQ(9); or (3) a decreased amount of ubiquinone.
We extracted the endogenous quinones from wildtype (N2) and clk-1 mitochondria, replenished them with exogenous ubiquinones, and measured ETC activities.
Replenishment of extracted mutant and wildtype mitochondria resulted in equal enzymatic activities for complexes I-III and II-III ETC assays. Blue native gels showed that supercomplex formation was indistinguishable between clk-1 and N2. The addition of a pentane extract from clk-1 mitochondria containing DMQ(9) to wildtype mitochondria specifically inhibited complexes I-III activity. UQ in clk-1 mitochondria was oxidized compared to N2.
Our results show that no measurable intrinsic ETC defect exists in clk-1 mitochondria. The data indicate that DMQ(9) specifically inhibits electron transfer from complex I to ubiquinone.