title: Purification and characterization of tissue kallikrein-like proteinases from the black sea bass (Centropristis striata) and the southern frog (Rana berlandieri).
abstract: Serine proteinases were isolated from the pyloric caeca of the black sea bass (Centropristis striata) and the pancreas of the Southern frog (Rana berlandieri) and were purified to apparent homogeneity by aprotinin affinity column chromatography, reverse phase high performance liquid chromatography and gel filtration FPLC liquid chromatography to produce products with molecular masses of approximately 27,000 Da and isoelectric points from 4.2 to 5.0. Both enzymes were kallikrein-like and were bound by diisopropylfluorophosphate; had pH optima from 9 to 10; showed high specificity for the hydrolysis of arginine peptide bonds and low to moderate affinity for lysine bonds at the P1 substrate recognition sites; were inhibited by aprotinin, benzamidine, leupeptin, and soybean trypsin inhibitor; generated kinin from kininogen and were highly stable at room temperature. Differences between the enzymes were observed relative to their hydrophobicities, substrate specificities, stabilities at acidic pHs in the presence and absence of calcium, and the amounts of kinin generated from kininogen. Many of the fish trypsins, previously identified as anionic trypsins, may actually be more kallikrein-like.
authors.affiliation: National Marine Fisheries Service, Southeast Fisheries Science Center, Charleston Laboratory, SC 29422, USA.
authors.forename: G P
authors.initials: GP
authors.lastname: Richards
authors.forename: J
authors.initials: J
authors.lastname: Chao
authors.forename: P
authors.initials: P
authors.lastname: Chung
authors.forename: L
authors.initials: L
authors.lastname: Chao
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D001492:Bass / Q000378:metabolism*; D002118:Calcium / Q000494:pharmacology; D002846:Chromatography, Affinity; D002851:Chromatography, High Pressure Liquid; D004591:Electrophoresis, Polyacrylamide Gel; D004795:Enzyme Stability / Q000187:drug effects; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D007525:Isoelectric Focusing; D007610:Kallikreins / Q000737:chemistry / Q000378:metabolism; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology*; D011708:Pylorus / Q000201:enzymology*; D011898:Ranidae / Q000378:metabolism*; D012697:Serine Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D013379:Substrate Specificity; D020840:Tissue Kallikreins
chemical_list: D007610:Kallikreins; D012697:Serine Endopeptidases; D020840:Tissue Kallikreins; D002118:Calcium