Lr. Dick et al., MECHANISTIC STUDIES ON THE INACTIVATION OF THE PROTEASOME BY LACTACYSTIN IN CULTURED-CELLS, The Journal of biological chemistry, 272(1), 1997, pp. 182-188
The natural product lactacystin exerts its cellular antiproliferative
effects through a mechanism involving acylation and inhibition of the
proteasome, a cytosolic proteinase complex that is an essential compon
ent of the ubiquitin-proteasome pathway for intracellular protein degr
adation, In vitro, lactacystin does not react with the proteasome; rat
her, it undergoes a spontaneous conversion (lactonization) to the acti
ve proteasome inhibitor, clasto-lactacystin beta-lactone. We show here
that when the beta-lactone is added to mammalian cells in culture, it
rapidly enters the cells, where it can react with the sulfhydryl of g
lutathione to form a thioester adduct that is both structurally and fu
nctionally analogous to lactacystin, We call this adduct lactathione,
and like lactacystin, it does not react with the proteasome, but can u
ndergo lactonization to yield back the active beta-lactonee, We have s
tudied the kinetics of this reaction under appropriate in vitro condit
ions as well as the kinetics of lactathione accumulation and proteasom
e inhibition in cells treated with lactacystin or beta-lactone, The re
sults indicate that only the beta-lactone (not lactacystin) can enter
cells and suggest that the formation of lactathione serves to concentr
ate the inhibitor inside cells, providing a reservoir for prolonged re
lease of the active beta-lactone.