Purification and biochemical characterization of phospholipase A2 from dromedary pancreas.
Dromedary pancreatic PLA2 (DrPLA2) was purified from delipidated pancreases. Pure protein was obtained after heat and acidic treatment (70 degrees C; pH 3.0), precipitation by ammonium sulphate and ethanol respectively, followed by sequential column chromatographies on Sephadex G-50, MonoS Sepharose, MonoQ Sepharose and C-8 reverse phase high pressure liquid chromatography. Purified DrPLA2, which is not glycosylated protein, was found to be monomeric protein with a molecular mass of 13748.55 Da. A specific activity of 600 U/mg for purified DrPLA2 was measured at optimal conditions (pH 8.0 and 37 degrees C) in the presence of 3 mM NaTDC and 7 mM CaCl(2) using PC as substrate. The sequence of the first fourteen amino-acid residues at the N-terminal extremity of DrPLA2 was determined by automatic Edman degradation. One single sequence was obtained and shows a close similarity with all other known pancreatic secreted phospholipases A2.
Bacha
Abir Ben
AB
Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, 3038 Sfax, Tunisia.
Gargouri
Youssef
Y
Bezzine
Sofiane
S
Mejdoub
Hafedh
H
D000818:Animals; D002162:Camelus; D002852:Chromatography, Ion Exchange; D004591:Electrophoresis, Polyacrylamide Gel; D006863:Hydrogen-Ion Concentration; D010179:Pancreas / Q000201:enzymology*; D010741:Phospholipases A / Q000302:isolation & purification* / Q000378:metabolism*; D054467:Phospholipases A2; D013696:Temperature
D010741:Phospholipases A; D054467:Phospholipases A2