Protein degradation by ERp72 from rat and mouse liver endoplasmic reticulum.
The endoplasmic reticulum (ER) resident protein, ER60, is a member of the protein disulfide-isomerase family and contains two copies of the internal thioredoxin motif, CGHC. Previously, ER60 was identified as a cysteine protease and named ER-60 protease (Urade, R., Nasu, M., Moriyama, T., Wada, K., and Kito, M. (1992) J. Biol. Chem. 267, 15152-15159; Urade, R., and Kito, M. (1992) FEBS Lett. 312, 83-86). Here, ERp72, the other member of the protein disulfide-isomerase family containing three CGHC motifs, was isolated from ER of rat and mouse livers through four sequential chromatographies on DEAE-Toyopearl 650, AF-heparin Toyopearl 650M, and TSK gel G3000SW twice. The purified rat protein was found to be homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, not being contaminated by ER-60 protease, as judged on immunoblot analysis using an anti-ER-60 protease antibody. The partial amino acid sequence of rat ERp72 was 93% homologous to that of mouse ERp72. The purified rat ERp72 degraded other ER resident proteins such as protein disulfide-isomerase and calreticulin. The purified mouse ERp72 also degraded those proteins. Though rat ERp72 did not basically require Ca2+ for the reaction, the degradation of protein disulfide-isomerase was enhanced, but the degradation of calreticulin was inhibited in the presence of Ca2+. The proteolytic activity of rat ERp72 was inhibited by cysteine protease inhibitors. Its sensitivity to protease inhibitors was the same as that of ER-60 protease. In addition, the proteolytic activity of rat ERp72 was inhibited by acidic phospholipids, also similar to ER-60 protease. Therefore, we propose that ERp72 be named ER-72 protease.
Urade
R
R
Research Institute for Food Science, Kyoto University, Japan.
Takenaka
Y
Y
Kito
M
M
D000595:Amino Acid Sequence; D000818:Animals; D010450:Endopeptidases / Q000378:metabolism*; D004721:Endoplasmic Reticulum / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D008099:Liver / Q000378:metabolism*; D008297:Male; D008562:Membrane Glycoproteins / Q000037:antagonists & inhibitors / Q000302:isolation & purification / Q000378:metabolism*; D051379:Mice; D008969:Molecular Sequence Data; D010743:Phospholipids / Q000494:pharmacology; D011480:Protease Inhibitors / Q000494:pharmacology; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley
D008562:Membrane Glycoproteins; D010743:Phospholipids; D011480:Protease Inhibitors; D011506:Proteins; C062477:endoplasmic reticulum glycoprotein p72; D010450:Endopeptidases