Purification of turkey pancreatic phospholipase A2.
Turkey pancreatic phospholipase (TPP) has been purified from delipidated pancreases. The purification included ammonium sulfate fractionation, acidic (pH 5) treatment, followed by sequencial column chromatographies on DEAE-cellulose, Sephadex G-75, and reverse phase high pressure liquid chromatography. The purified enzyme was found to be a monomeric protein with molecular mass of 14 kDa. The optimal activity was measured at pH 8 and 37 degrees C using egg yolk emulsion as substrate. Our results show that the enzyme (TPP) was not stable for 1 h at 60 degrees C, and that bile salt and Ca2+ were required for the expression of the purified enzyme. The sequence of the N-terminal amino acids of the purified enzyme shows a very close similarity between TPP and all other known pancreatic phospholipases.
Laboratory of Biochemistry, ENIS Soukra Road, 3038 Sfax, Tunisia.
Riadh
R
Ben Salah
Nacim
N
Zouari
Joseph
J
Reinbolt
Hafedh
H
Mejdoub
D000595:Amino Acid Sequence; D000645:Ammonium Sulfate; D000818:Animals; D001647:Bile Acids and Salts; D002118:Calcium; D002845:Chromatography; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010179:Pancreas / Q000201:enzymology*; D010741:Phospholipases A / Q000302:isolation & purification*; D054467:Phospholipases A2; D013696:Temperature; D014422:Turkeys
D001647:Bile Acids and Salts; D010741:Phospholipases A; D054467:Phospholipases A2; D000645:Ammonium Sulfate; D002118:Calcium