The echinocandins are membrane-anchored, cyclic lipopeptides (CLPs) with antifungal activity due to their ability to inhibit a glucan synthase located in the plasma membrane of fungi such as Candida albicans. A hydrophobic tail of an echinocandin CLP inserts into a membrane, placing a six-amino acid cyclic peptide near the membrane surface. Because processes critical for the function of the electron transfer complexes of mitochondria, such as proton uptake and release, take place near the surface of the membrane, we have tested the ability of two echinocandin CLPs, caspofungin and micafungin, to affect the activity of electron transfer complexes in isolated mammalian mitochondria. Indeed, caspofungin and micafungin both inhibit whole chain electron transfer in isolated mitochondria at low micromolar concentrations. The effects of the CLPs are fully reversible, in some cases simply via the addition of bovine serum albumin to bind the CLPs via their hydrophobic tails. Each CLP affects more than one complex, but they still exhibit specificity of action. Only caspofungin inhibits complex I, and the CLP inhibits liver but not heart complex I. Both CLPs inhibit heart and liver complex III. Caspofungin inhibits complex IV activity, while, remarkably, micafungin stimulates complex IV activity nearly 3-fold. Using a variety of assays, we have developed initial hypotheses for the mechanisms by which caspofungin and micafungin alter the activities of complexes IV and III. The dication caspofungin partially inhibits cytochrome c binding at the low-affinity binding site of complex IV, while it also appears to inhibit the release of protons from the outer surface of the complex, similar to Zn(2+). Anionic micafungin appears to stimulate complex IV activity by enhancing the transfer of protons to the O2 reduction site. For complex III, we hypothesize that each CLP binds to the cytochrome b subunit and the Fe-S subunit to inhibit the required rotational movement of the latter.