LOCATION AND CHARACTERIZATION OF AMINOPEPTIDASE-N IN LACTOCOCCUS-LACTIS SUBSP CREMORIS HP

One single, cytosolic aminopeptidase (AP N, EC 3.4.11.2) is found to be responsible for both leucyl- (leucylAP) and lysylaminopeptidase (lysylAP) activity detectable with whole cells of Lactococcus lactis subsp. cremoris strain HP. The existence of a cell-envelope-located form of this enzyme could be excluded. No restriction on the activity of the enzyme is imposed by the cell membrane if leucine-p-nitroanilide is used as the substrate; with lysine-p-nitroanilide the activity is highly cryptic. The enzyme has been purified and characterized. It is a metalloaminopeptidase with a molecular mass of 95 kDa. Co2+ appears to be the most potent ion to (re)activate the enzyme; Zn2+ and Mn2+ are less effective. The AP N releases the positively charged amino acids and several uncharged (including proline) from the N-terminus. Ammonium salts affect the preference of the enzyme with respect to the N-terminal residue. A preferential interaction of the ammonium ion with an essential cation binding site seems to be responsible for the inhibition of lysylAP activity.