Trypsin from the viscera of Bogue (Boops boops): isolation and characterisation.
Trypsin from the viscera of Bogue (Boops boops) was purified to homogeneity by precipitation with ammonium sulphate, Sephadex G-100 gel filtration and Mono Q-Sepharose anion exchange chromatography, with an 8.5-fold increase in specific activity and 36% recovery. The molecular weight of the purified enzyme was estimated to be 23 kDa by SDS-PAGE and size exclusion chromatography. The purified trypsin appeared as a single band on native-PAGE and zymography staining. The purified enzyme showed esterase-specific activity on N-α-benzoyl-L-arginine ethyl ester (BAEE) and amidase activity on N-α-benzoyl-DL-arginine-p-nitroanilide (BAPNA). The optimum pH and temperature for the enzyme activity, after 10 min incubation, were pH 9.0 and 55°C, respectively, using BAPNA as a substrate. The trypsin kinetic constants Km and kcat on BAPNA were 0.13 mM and 1.56 s(-1), respectively, while the catalytic efficiency kcat/Km was 12 s(-1) mM(-1). Biochemical characterisation of B. boops trypsin showed that this enzyme can be used as a possible biotechnological tool in the fish processing and food industries.
Barkia
Ahmed
A
Laboratoire de Génie Enzymatique et de Microbiologie, Ecole Nationale d'Ingénieurs de Sfax, B.P. 1173, 3038, Sfax, Tunisia.
Bougatef
Ali
A
Nasri
Rim
R
Fetoui
Emna
E
Balti
Rafik
R
Nasri
Moncef
M
D000581:Amidohydrolases / Q000378:metabolism; D000818:Animals; D017756:Aquaculture / Q000379:methods; D001120:Arginine / Q000031:analogs & derivatives; D001586:Benzoylarginine Nitroanilide; D002850:Chromatography, Gel; D002852:Chromatography, Ion Exchange; D003911:Dextrans; D004950:Esterases / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010473:Perciformes / Q000378:metabolism*; D013696:Temperature; D014357:Trypsin / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism*; D014781:Viscera / Q000201:enzymology*
D003911:Dextrans; C025614:sephadex; D001586:Benzoylarginine Nitroanilide; D001120:Arginine; C009727:benzoylarginine ethyl ester; D004950:Esterases; D014357:Trypsin; D000581:Amidohydrolases; C019665:amidase