Identification of latent procathepsins B and L in microsomal lumen: characterization of enzymatic activation and proteolytic processing in vitro.
Procathepsins B and L in the hepatic endoplasmic lumen were identified as having a molecular weight of 39,000 by immunoblot analysis. The proenzymes were then purified to remove the mature enzymes by concanavalin A-Sepharose chromatography. The concanavalin A-adsorbed fractions containing the proenzymes showed no appreciable activities of cathepsins B and L. When those fractions were incubated at pH 3.0, the enzymatic activities markedly increased: the activities of cathepsins B and L after 36 h incubation were 60 and 210 times those of the controls, respectively. Immunoblot analysis showed that after 36 h incubation the proenzymes disappeared and the mature enzymes increased. Thus the proenzymes were processed to the mature enzymes under acidic conditions of pH 3.0. The marked increases of enzymatic activities and the conversion of the proenzymes to the mature forms were completely blocked with pepstatin, which is a potent inhibitor of aspartic proteases. The results strongly suggested that a processing protease for procathepsins B and L might be cathepsin D, a major lysosomal aspartic protease. Indeed, lysosomal cathepsin D could convert microsomal procathepsin B to the mature enzyme in vitro. Therefore, procathepsins B and L seem first to be synthesized as enzymatically inactive forms in endoplasmic reticulum and successively may be converted into active forms by cathepsin D in lysosomal compartments.
Nishimura
Y
Y
Department of Physiological Chemistry, Faculty of Pharmaceutical Sciences, Kyushu University, Fukuoka, Japan.
Kawabata
T
T
Kato
K
K
D000818:Animals; D002401:Cathepsin B / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D056668:Cathepsin L; D002403:Cathepsins / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D003546:Cysteine Endopeptidases; D010450:Endopeptidases; D004789:Enzyme Activation; D004792:Enzyme Precursors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D007700:Kinetics; D008297:Male; D008862:Microsomes, Liver / Q000201:enzymology*; D008970:Molecular Weight; D011499:Protein Processing, Post-Translational; D051381:Rats; D011919:Rats, Inbred Strains
D004792:Enzyme Precursors; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D002401:Cathepsin B; D056668:Cathepsin L; C534280:Ctsl protein, rat