Pancreatic bile salt dependent lipase from cod (Gadus morhua): purification and properties.
The enzymatic basis for cod digestive lipolysis has been investigated. Lipase activity was found in aqueous extracts from pyloric caeca as well as in pancreatic tissue surrounding the caeca and the bile duct. A bile salt-dependent lipase (BSDL) was purified from either defatted powder of cod pyloric caeca or aqueous pancreatic extracts by combined affinity chromatography on cholate-Sepharose and gel filtration on Sephacryl S-200 HR. By SDS-PAGE analysis the molecular weight of purified cod BSDL was estimated to 60 kDa. The enzyme was totally dependent on bile salts for hydrolysis of insoluble fatty acid esters. Antiserum raised against purified cod BSDL reacted specifically with selected mammalian pancreatic BSDLs by Western blot analysis. Results presented in this paper strongly suggest that the bile salt-dependent lipase is the only pancreatic enzyme involved in lipid digestion in cod. The enzyme has been characterized and compared to human pancreatic BSDL with respect to substrate specificity, temperature- and pH-dependence and inhibitors. Both soluble and insoluble fatty acid esters were hydrolysed and the enzyme was 1,3-specific in hydrolysis of triolein. The enzyme was inhibited by di-isopropyl fluorophosphate and phenyl boronic acid, but not significantly by phenyl methyl sulfonyl fluoride. The cod BSDL is probably homologous to mammalian pancreatic BSDLs.
Gjellesvik
D R
DR
Department of Biochemistry, University of Bergen, Norway.
Lombardo
D
D
Walther
B T
BT
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D001647:Bile Acids and Salts / Q000378:metabolism*; D015153:Blotting, Western; D001897:Boronic Acids / Q000494:pharmacology; D002384:Catalysis; D005399:Fishes / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007531:Isoflurophate / Q000494:pharmacology; D007700:Kinetics; D008049:Lipase / Q000037:antagonists & inhibitors / Q000302:isolation & purification*; D010179:Pancreas / Q000201:enzymology*; D013379:Substrate Specificity; D014304:Triolein / Q000378:metabolism
D000596:Amino Acids; D001647:Bile Acids and Salts; D001897:Boronic Acids; D014304:Triolein; D007531:Isoflurophate; D008049:Lipase; C010686:benzeneboronic acid