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Advisor(s)
Abstract(s)
An intracellular hydrolase able to cleave Image -lysine-p-nitroanilide was purified from Lactobacillus plantarum strain ESB5004 via two steps of precipitation with ammonium sulfate (at 30 and 50% (w/v)), followed by hydrophobic interaction and ion-exchange chromatographies. The aminopeptidase was purified up to 11-fold, with a final yield of ca. 1%. Its native molecular weight is ca. 70 kDa, and it is apparently composed of two subunits, the molecular weight of which is 34 kDa. The enzyme was assayed using a wide variety of p-nitroanilide (pNA) derivatives as substrates: it hydrolyzed preferentially pNA adducts of hydrophobic and basic amino acid residues; no hydrolysis was in particular observed of Glu-pNA, Gly-pNA or Pro-pNA. The enzyme activity was removed by the metal-chelating agent EDTA, thus suggesting that it is a metallo-enzyme; however, the EDTA-inhibited enzyme was reactivated in the presence of Co2+. Optimal aminopeptidase activity was obtained at 28 °C (pH 7.0) and pH 6.5 (37 °C). The enzyme was inhibited by 10 mM CaCl2 or MgCl2.
Description
Keywords
Enzyme Isolation Activity Lactic acid bacteria Dairy food
Pedagogical Context
Citation
MACEDO, Angela C. ; TAVARES, Tânia G. ; MALCATA, F. Xavier - Purification and characterization of an intracellular aminopeptidase from a wild strain of Lactobacillus plantarum isolated from traditional Serra da Estrela cheese. Enzyme and Microbial Technology. ISSN: 0141-0229. Vol. 32, n.º 1 (2003) p. 41-48
Publisher
Elsevier