A new aminopeptidase was isolated from the biomass of the flagellate Astasia longa by precipitation with ammonium sulfate, gel filtration, and affinity chromatography on Arginine-Silochrome in 41% yield and with purification degree 490. The enzyme is irreversible inhibited by mercury chloride, EDTA, o-phenanthroline and, partially, bestatin and zinc chloride. It has an optimum pH 8.5 toward the hydrolysis of a synthetic chromogenic substrate Ala-pNA. The enzyme molecular mass is 45 kDa, isoelectric point 5.5, and temperature optimum 45 degrees C. The enzyme most effectively hydrolyzes p-nitroanilides of alanine, arginine, and leucine; it is classified as metalloaminopeptidase.
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A new aminopeptidase was isolated from the biomass of the flagellate Astasia longa by precipitation with ammonium sulfate, gel filtration, and affinity chromatography on Arginine-Silochrome in 41% yield and with purification degree 490. The enzyme is irreversible inhibited by mercury chloride, EDTA, o-phenanthroline and, partially, bestatin and zinc chloride. It has an optimum pH 8.
View Article and Find Full Text PDFPlasma membranes (PM) were prepared by discontinuous density gradient centrifugation of crude nuclear fractions from 6 rat livers. These "nuclear" PM (PM-n) were 15-fold enriched in plasma membrane marker enzymes and contained an endopeptidase activity degrading azocasein at pH 7. To get larger amounts of plasma membranes, microsomal fractions obtained in large scale subcellular fractionations were subjected to continuous gradient zonal centrifugation.
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