Bacillus kaustophilus leucine aminopeptidase (bkLAP) was sensitive to oxidative damage by hydrogen peroxide. To improve its oxidative stability, the oxidation-sensitive methionine residues in the enzyme were replaced with leucine by site-directed mutagenesis. The variants, each with an apparent molecular mass of approximately 54 kDa, were overexpressed in recombinant Escherichia coli M15 cells and purified to homogeneity by nickel-chelate chromatography. The specific activity for M282L, M285L, M289L and M321L decreased by more than 43%, while M400L, M426L, M445L, and M485L showed 191, 79, 313, and 103%, respectively, higher activity than the wild-type enzyme. Although the mutations did not cause significant changes in the K(m) value, more than 67.8% increase in the value of k(cat)/K(m) was observed in the M400L, M426L, M445L and M485L. In the presence of 50 mM H2O2, most variants were more stable with respect to the wild-type enzyme, indicating that the oxidative stability of the enzyme can be improved by engineering the methionine residues.