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1. Psychosine (beta-galactosylsphingosine) is the toxic agent in Krabbe's disease (globoid cells leukodystrophy). It inhibits purified bovine heart mitochondrial cytochrome c oxidase; there is a rapid phase of inhibition (complete within 10-15 s) and a slower phase (complete within 10-15 min). Both phases are also seen in rat liver mitochondria. IC50 is about 200 microM psychosine in the purified enzyme and less than 20 microM in mitochondria. Psychosine inhibition is due to binding to cytochrome oxidase, not cytochrome c. 2. Bovine heart submitochondrial particles show inhibition similar to rat liver mitochondria. However, although proteoliposomes containing bovine heart cytochrome oxidase show an identical fast phase, they have no noticeable slow phase of inhibition. Addition of phospholipid liposomes to submitochondrial particles relieved the majority of psychosine inhibition, consistent with the removal of those molecules binding in the slow phase. Psychosine can inhibit cytochrome oxidase molecules facing in either direction in proteoliposomes and submitochondrial particles, suggesting that it can rapidly interact with both sides of a membrane when added externally. 3. At high ionic strength, the presence of psychosine decreases the Vmax. of cytochrome oxidase with little effect on the Km for cytochrome c. This non-competitive inhibition suggests that the psychosine-enzyme complex is kinetically inactive and not labile over the time course of the assay. Psychosine does not inhibit the reduction of haem a or haem a3 by artificial electron donors, but does inhibit the reduction of haem a by cytochrome c.

Kinetics of inhibition of purified and mitochondrial cytochrome c oxidase by psychosine (β-galactosylsphingosine)