Characterization of a triglyceride-lipase and its natural substrates from crustacean hepatopancreas

PASQUEVICH M. Y.; DREON M. S.; LAVARÍAS S.; HERAS H.

San Miguel de Tucumán

Congreso; XLV Reunion Anual - Sociedad Argentina de Investigación en Bioquímica y Biología Molecular; 2009

Sociedad Argentina de Investigaciones Bioquímicas

The aim of this work was to purify and characterize a citosolic triglyceride-lipase (lipase) activity and its natural substrates from the hepatopancreas of Macrobrachium borellii. Purification asfollowed by a zymographic assay and the esterase activity was confirmed by the hydrolysis of "C-TG. The optimal temperature and pH values were 36°C and 8.0, respectively. The kineticcharacterization using pNPP as substrate showed a Michaelis-Menten behavior with a Vmax=3.4x101 erization of a lipase and its natural substrates in crustaceans.umol.min-1 .mg- 1 and a Km=1.63 mM. A MW of 72 KDa was estimated by PAGE. The TGs speciesfrom M. borellii hepatopancreas were studied by silver nitrate chromatography allowing the separation of several groups of increasing Rf hereafter named group I-IV. Fatty acid (FA)composition of each TG group was analyzed by capillary GC. Group I was the major PUFA-containing one (48.5%) followed by group I1 (45.5%). Group I11 was dominated by monounsaturated FA (58.3%), while group IV had both monounsaturated and saturatedFA(50.0% and45.6%, respectively). Finally substrate specificity of lipase was determined bycompetence of Group I and different non-radiolabeled TGs on the rate of hydrolysis of labeled triolein. The enzyme showed higher specificity toward TG PUFA-containing species than towardsaturated TGs. This is the first purification and characterization of a lipase and itsnatural substrates in crustaceans.