Characterization of a neutral aminoacyl-peptide hydrolase from Naegleria fowleri.
An intracellular alpha-aminoacyl-peptide hydrolase (EC 3.4.11.-) from Naegleria fowleri nN68 (ATCC 30894) has been characterized. The enzyme preparation hydrolyzed phenylalanyl-, tyrosyl-, leucyl-, arginyl-, alanyl-, tryptophanyl-, histidyl-, methionyl-, and lysyl-naphthylamide but not benzoylleucyl-, leucylglycyl-, glycylprolylleucyl-, glycyl-, threonyl-, aspartyl-, or glutamyl-naphthylamide. The aminopeptidase activity was inhibited by the cysteine-protease inhibitors--hydroxymercuribenzoate, chloromercurisulfate, and iodoacetate--by the aminopeptidase inhibitors--bestatin and trans-epoxysuccinyl-leucyl-agmatine--by an inhibitor of soluble alanyl aminopeptidase EC 3.4.11.14, puromycin, and by the metalloprotease inhibitor, o-phenanthroline. The exopeptidase activity was not inhibited by the chelator, ethylenediaminetetraacetate, or the serine-protease inhibitor, phenylmethylsulfonylfluoride. The pH optimum of the exopeptidase was between 7.0 and 8.0. Enzyme activity was stable at 55 degrees C for 30 min, but all activity was lost after 15 min at 80 degrees C. Enzyme activity was inhibited by 100 microM HgCl2 and CdCl2 but not by 1 mM CoCl2, CuCl2, MnCl2, NiCl2, FeCl3, or ZnCl2. Enzyme activity was inhibited by 0.1% sodium dodecyl sulfate but not by 0.2% Brij 35, Tween 20, Tween 80, or Triton X-100.
D000656:Amoeba / Q000201:enzymology*; D000818:Animals; D003902:Detergents / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007930:Leucine / Q000031:analogs & derivatives / Q000378:metabolism; D007931:Leucyl Aminopeptidase / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009994:Osmolar Concentration; D013379:Substrate Specificity; D013696:Temperature
D003902:Detergents; C040135:leucine-beta-naphthylamide; D007931:Leucyl Aminopeptidase; D007930:Leucine