TAILIEUCHUNG - Báo cáo Y học: Myristyl and palmityl acylation of pI 5.1 carboxylesterase from porcine intestine and liver Tissue and subcellular distribution

Immunoblotting analyses revealed the presence of carb-oxylesterase in theporcine small intestine, liver, submaxillary andparotidglands, kidney cortex, lungs and cerebral cortex. In the intestinal mucosa, the pI enzyme was detected in several subcellular fractions including the microvillar frac-tion. Both fatty monoacylated and diacylated monomeric (F1), trimeric (F3) and tetrameric (F4) forms of the intestinal protein were puri®ed here for the ®rst time by performing hydrophobic chromatography and gel ®ltration | Eur. J. Biochem. 269 1109-1117 2002 FEBS 2002 Myristyl and palmityl acylation of pl carboxylesterase from porcine intestine and liver Tissue and subcellular distribution Sylvie Smialowski-Fleter Andre Moulin Josette Perrier and Antoine Puigserver Institut Mediterraneen de Recherche en Nutrition UMR-INRA Faculte des Sciences et Techniques de St-Jerome Marseille France Immunoblotting analyses revealed the presence of carboxylesterase in the porcine small intestine liver submaxillary and parotid glands kidney cortex lungs and cerebral cortex. In the intestinal mucosa the pI enzyme was detected in several subcellular fractions including the microvillar fraction. Both fatty monoacylated and diacylated monomeric F1 trimeric F3 and tetrameric F4 forms of the intestinal protein were purified here for the first time by performing hydrophobic chromatography and gel filtration. The molecular mass of these three enzymatic forms was estimated to be 60 180 and 240 kDa respectively based on sizeexclusion chromatography and SDS PAGE analysis. The existence of a covalent attachment linking palmitate and myristate to porcine intestinal carboxylesterase PICE which was suggested by the results of gas-liquid chromatography GLC experiments in which the fatty acids resulting from alkali treatment of the protein forms were isolated was confirmed here by the fact that 3H palmitic and 3H myristic acids were incorporated into porcine enterocytes and hepatocytes in cell primary cultures. Besides these two main fatty acids the presence of oleic stearic and arachidonic acids was also detected by GLC and further confirmed by performing radioactivity counts on the 3H-labelled PICE forms after an immunoprecipitation procedure using specific polyclonal antibodies followed by a SDS PAGE separation step. Unlike the F1 and F4 forms which were both myristoylated and palmitoylated the F3 form was only palmitoylated. The monomeric trimeric and tetrameric forms of PICE were all able to hydrolyse .

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