Separation and properties of three forms of cathepsin H-like cysteine proteinase from rat spleen.
Three forms of cathepsin H-like cysteine proteinase were purified from rat spleen by a method involving acid treatment and chromatography on pepstatin-Sepharose, Sephadex G-75, DEAE-Sephacel, CM-Toyopearl, and concanavalin A-Sepharose. The final preparations of these forms all migrated as single protein bands on polyacrylamide gel electrophoresis with and without sodium dodecyl sulfate (SDS). The molecular weights of the three forms were estimated to be 28,000 (form I), 26,000 (form II), and 22,000 (form III). The optimal pH was 6.5 for forms I and III and was 7.0 for form II with L-leucine 2-naphthylamide (Leu-NA) or with alpha-N-benzoyl-DL-arginine 2-naphthylamide (BANA). All of the forms consisted of two major species having isoelectric points of 7.1 and 6.5 on isoelectric focusing gels. They were all stable when incubated at pH values between 5.0 and 9.0 for 1 h at 22 degrees C. They were strongly inhibited by iodoacetic acid and E-64, but not by metal ions or pepstatin. Form III was not affected by leupeptin, chymostatin, antipain or elastatinal, which gave essentially complete inhibition of cathepsin B purified from rat spleen. Forms I and II were slightly inhibited by these compounds at the same concentrations. The properties of these forms were compared with those of the known enzymes cathepsin H and BANA-hydrolase.
D000818:Animals; D056655:Cathepsin H; D002403:Cathepsins / Q000302:isolation & purification*; D002850:Chromatography, Gel; D003546:Cysteine Endopeptidases; D004591:Electrophoresis, Polyacrylamide Gel; D010450:Endopeptidases / Q000302:isolation & purification*; D004789:Enzyme Activation / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D011480:Protease Inhibitors / Q000494:pharmacology; D051381:Rats; D013154:Spleen / Q000201:enzymology*; D013438:Sulfhydryl Compounds / Q000494:pharmacology
D011480:Protease Inhibitors; D013438:Sulfhydryl Compounds; D002403:Cathepsins; D010450:Endopeptidases; D003546:Cysteine Endopeptidases; D056655:Cathepsin H; C534128:Ctsh protein, rat