Purification and biochemical characterization of a turkey preduodenal esterase.
A preduodenal esterase was purified to homogeneity from turkey (Meleagris gallopavo) pharyngial tissue. Pure turkey pregastric esterase (TPrE) was obtained after anion exchange chromatography (DEAE-cellulose), Sephacryl S-200 gel filtration, anion exchange chromatography (Mono-Q sepharose) and affinity chromatography (Blue-Gel Affi Gel). The pure enzyme has an apparent molecular mass of 50 kDa, as determined by SDS/PAGE analysis. The optimum pH and temperature for enzyme activity using tributyrin as substrate were 8.5 and 48 degrees C, respectively. Under these conditions, the specific activity measured was 650 U/mg. No significant lipolytic activity was found when was tested on triolein or liprocil as substrates or with monolayer dicaprin with TPrE. In contrast, TPrE displayed a maximal activities of 800, 680 and 520 U/mg with vinyl acetate, vinyl propionate and vinyl butyrate, respectively. This enzyme retained 75% of its maximal activity when incubated for 30 min at pH 4 and 50 degrees C, but it was completely inactivated after an incubation for 10 min at 60 degrees C. The TPrE N-terminal amino acid sequence showed similarities to the N-terminal sequence of a thioesterase from mallard duck, but no similarity with known preduodenal lipases was found.
Cherif
Slim
S
Laboratoire de Biochimie et de Génie Enzymatique des Lipases, ENIS route de Soukra, 3038 Sfax-Tunisia.
Miled
Nabil
N
Kammoun
Maher
M
Mejdoub
Hafedh
H
Gargouri
Youssef
Y
D000595:Amino Acid Sequence; D000818:Animals; D001647:Bile Acids and Salts / Q000494:pharmacology; D002846:Chromatography, Affinity; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003426:Crop, Avian / Q000201:enzymology*; D004795:Enzyme Stability; D004950:Esterases / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D013696:Temperature; D014422:Turkeys / Q000378:metabolism*
D001647:Bile Acids and Salts; D004950:Esterases