Separation of cathepsin A-like enzyme and the proteasome: evidence that lactacystin/beta-lactone is not a specific inhibitor of the proteasome.
Previous studies have described a human platelet cathepsin A-like enzyme with a number of similarities to the "acidic" and "neutral" chymotrypsin-like activities of the proteasome. This includes its strong inhibition by the highly specific proteasome inhibitor Lactacystin/beta-lactone, suggesting that either the Cbz-Phe-Ala-hydrolyzing activity attributed to cathepsin A was due to the chymotrypsin-like activity of the proteasome or that lactacystin was not a specific inhibitor of the proteasome. In the present study we discard the first possibility on the basis of the following findings: (a) human platelet cathepsin A, unlike proteasome, binds to concanavalin A, and does not bind to Heparin-Sepharose at pH 7.4; (b) neither the chymotrypsin-like activity of the proteasome, nor proteasome antigens are detected in the cathepsin A preparation; (c) purified proteasome does not exhibit Cbz-Phe-Ala-hydrolyzing activity; (d) Z-lle-Glu-(Ot-Bu)Ala-leucinal (PSI), a compound that selectively inhibits the chymotrypsin-like activity of the proteasome at a concentration of 10 microM has no inhibitory effect on the carboxypeptidase activity of cathepsin A; (e) cathepsin A, free of the proteasome, is completely inhibited by micromolar concentrations of lactacystin/beta-lactone. It is therefore concluded that lactacystin/beta-lactone is not a specific inhibitor of the proteasome.
Ostrowska
H
H
Department of Instrumental Analysis, and Department of Oncology Medical Academy, Bialystok, Poland. halost@amb.ac.bialystok.pl
Wójcik
C
C
Wilk
S
S
Omura
S
S
Kozlowski
L
L
Stoklosa
T
T
Worowski
K
K
Radziwon
P
P
D000111:Acetylcysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D001792:Blood Platelets / Q000201:enzymology* / Q000378:metabolism; D043402:Cathepsin A / Q000037:antagonists & inhibitors* / Q000302:isolation & purification; D045744:Cell Line, Tumor; D002846:Chromatography, Affinity; D003208:Concanavalin A / Q000378:metabolism; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology*; D006493:Heparin / Q000378:metabolism; D006801:Humans; D051379:Mice; D046988:Proteasome Endopeptidase Complex / Q000302:isolation & purification; D061988:Proteasome Inhibitors
D015853:Cysteine Proteinase Inhibitors; D061988:Proteasome Inhibitors; D003208:Concanavalin A; C067713:lactacystin; D006493:Heparin; D043402:Cathepsin A; D046988:Proteasome Endopeptidase Complex; D000111:Acetylcysteine