Presence of oxidized protein hydrolase in human cell lines, rat tissues, and human/rat plasma.
Oxidized protein hydrolase (OPH), an 80 kDa serine protease whose activity is inhibited by diisopropyl fluorophosphate (DFP), has been isolated from human erythrocytes [Fujino, T. et al. (1998) J. Biochem. 124, 1077-1085]. The presence of OPH in various biological samples was examined by enzyme-linked immunosorbent assay (ELISA) and immunoblotting using an anti-OPH antibody raised against OPH purified from human erythrocytes, and by [(3)H]DFP-labeling and successive SDS-PAGE/fluorography. Solubilized samples of human cell lines including K-562 cells, THP-1 cells and Jurkat cells, and rat tissues including brain, heart, liver, kidney, and testis, inhibited the anti-OPH antibody binding to OPH in ELISA. Immunoblotting of lysates of K-562 cells, THP-1 cells and Jurkat cells showed four immunoreactive protein bands including an 80 kDa protein. Immunoprecipitation of the [(3)H]DFP-labeled K-562 cell lysate and successive SDS-PAGE/fluorography showed the presence of only the 80 kDa DFP-reactive protein with OPH antigenic activity. The level of the 80 kDa immunoreactive protein in K-562 cells rose as the cells differentiated toward erythrocytes. Immunoblotting of human and rat plasma showed two immunoreactive protein bands, including the 80 kDa protein, and SDS-PAGE/fluorography of [(3)H]DFP-labeled rat and human plasma showed the presence of only the 80 kDa DFP-reactive protein. The results indicate that OPH is present in a wide variety of biological samples.
Fujino
T
T
School of Pharmacy, Tokyo University of Pharmacy and Life Science, Horinouchi, Hachioji, Tokyo 192-0392, Japan.
Tada
T
T
Hosaka
T
T
Beppu
M
M
Kikugawa
K
K
D000818:Animals; D001921:Brain / Q000201:enzymology; D002454:Cell Differentiation; D002460:Cell Line / Q000201:enzymology; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D004912:Erythrocytes / Q000166:cytology / Q000201:enzymology*; D005470:Fluorometry / Q000379:methods; D006801:Humans; D015151:Immunoblotting; D007668:Kidney / Q000201:enzymology; D008099:Liver / Q000201:enzymology; D008297:Male; D009206:Myocardium / Q000201:enzymology; D010084:Oxidation-Reduction; D051381:Rats; D012697:Serine Endopeptidases / Q000097:blood / Q000276:immunology / Q000378:metabolism*; D013737:Testis / Q000201:enzymology; D014316:Tritium
D014316:Tritium; D012697:Serine Endopeptidases