title: Purification and properties of elastolytic enzyme from Flavobacterium immotum.
abstract: Elastolytic enzyme was purified and crystallized from culture fluid of Flavobacterium immotum No. 9-35. The purified enzyme was homogeneous on polyacrylamide gel electrophoresis. The molecular weight was determined by Sephadex G-100 gel filtration to be 13,000. The isoelectric point was between pH 8.3 and 8.9. The optimum pH of the enzyme was 7.2 for elastolytic activity. The purified enzyme showed not only elastolytic activity, but also non-specific proteolytic activity against various other proteins. Milk-clotting activity was also observed. The enzyme did not act on keratin, collagen, or fourteen amino acid esters, including N-benzoyl-L-alanine methyl ester, N-benzoyl-L-arginine ethyl ester, and N-acetyl-L-tyrosine ethyl ester, which were typical substrates of pancreatic elastase [EC 3.4.21.11], trypsin [EC 3.4.21.4], and chymotrypsin [EC 3.4.21.1], respectively. However, the enzyme selectively hydrolyzed elastin when both elastin and albumin were present in the reaction mixture. The enzyme was inhibited by o-phenanthroline and various heavy metals such as cadmium, lead, zinc, and mercury. Various inhibitors, such as diisopropyl phosphofluoridate, tosyl-L-lysine chloromethyl ketone, tosyl-L-phenylalanine chloromethyl ketone, trypsin inhibitor, iodoacetamide, etc., had no effect on the elastolytic activity.
mesh_terms: D000818:Animals; D002417:Cattle; D002848:Chromatography, DEAE-Cellulose; D002852:Chromatography, Ion Exchange; D003460:Crystallization; D005417:Flavobacterium / Q000201:enzymology*; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007531:Isoflurophate / Q000494:pharmacology; D007700:Kinetics; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology; D010196:Pancreatic Elastase / Q000302:isolation & purification* / Q000378:metabolism; D013045:Species Specificity; D013329:Structure-Activity Relationship; D013696:Temperature; D013997:Time Factors
chemical_list: D007531:Isoflurophate; D010196:Pancreatic Elastase