MECHANISTIC STUDIES ON THE INACTIVATION OF THE PROTEASOME BY LACTACYSTIN - A CENTRAL ROLE FOR CLASTO-LACTACYSTIN BETA-LACTONE

Lactacystin is a Streptomyces metabolite that inhibits cell cycle prog ression and induces differentiation in a murine neuroblastoma cell lin e, The cellular target of lactacystin is the 20 S proteasome, also kno wn as the multicatalytic proteinase complex, an essential component of the ubiquitin-proteasome pathway for intracellular protein degradatio n, In aqueous solution at pH 8, lactacystin undergoes spontaneous hydr olysis to yield N-acetyl-L-cysteine and the inactive lactacystin analo g, clasto-lactacystin dihydroxy acid, We have studied the mechanism of lactacystin hydrolysis under these conditions and found that it proce eds exclusively through the intermediacy of the active lactacystin ana log, clasto-lactacystin beta-lactone, Conditions that stabilize lactac ystin (and thus prevent the transient accumulation of the intermediate beta-lactone) negate the ability of lactacystin to inactivate the pro teasome. Together these findings suggest that lactacystin acts as a pr ecursor for clasto-lactacystin beta-lactone and that the latter is the sole species that interacts with the proteasome.