Location and characterization of aminopeptidase N in Lactococcus lactis subsp. cremoris HP

Summary One single, cytosolic aminopeptidase (AP N, EC 3.4.11.2) is found to be responsible for both leucyl-(leucylAP) and lysylaminopeptidese (lysylAP) activity detectible 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.Trainee from the Laboratory School Friesland, Leeuwarden, The NetherlandsOffprint requests to: F. A. Exterkate