A rapid procedure for the large scale purification of elastase and cathepsin G from human sputum.
A procedure is described which permits the rapid isolation of large amounts of elastase and cathepsin G from purulent sputum. This procedure involves: (1) digestion of sputum with DNase, (2) extraction of the insoluble residue that remains with 1 M NaCl, pH 8, (3) affinity chromatography on Sepharose-bound Trasylol, and (4) separation of the two enzymes by chromatogrphy on CM-Sephadex. Starting with 500 g of sputum it was possible to isolate 175 mg of each of these two enzymes within 7 to 10 days. Active site titration indicated both enzymes to be at least 97% pure. Disc gel electrophoresis in the presence and absence of SDS and amino acid sequence of the N-terminal region support the conclusion that the elastase and cathepsin G isolated from sputum are identical to the same enzymes isolated directly from the leukocytes of human blood.
D000595:Amino Acid Sequence; D001665:Binding Sites; D002403:Cathepsins / Q000302:isolation & purification*; D002846:Chromatography, Affinity / Q000379:methods; D002850:Chromatography, Gel / Q000379:methods; D003550:Cystic Fibrosis / Q000201:enzymology*; D003851:Deoxyribonucleases; D004589:Electrophoresis, Disc; D006801:Humans; D007962:Leukocytes / Q000201:enzymology; D010196:Pancreatic Elastase / Q000302:isolation & purification*; D013183:Sputum / Q000201:enzymology*; D013492:Suppuration
D003851:Deoxyribonucleases; D002403:Cathepsins; D010196:Pancreatic Elastase