Protease inhibitors from the Kunitz family (I3 in MEROPS) of 20-25 kDa are widely distributed in plant kingdom. They share a conserved b-trefoil fold in which a set of variable loops forms reactive centers for the interaction with target proteases. The majority of the Kunitz inhibitors are targeting serine proteases; however, inhibitors of cysteine and aspartic proteases have also been described. PDI (potato cathepsin D inhibitor) from the Kunitz family is a wound inducible plant defense protein against insect herbivores and pathogens. PDI is a unique bifunctional inhibitor of serine proteases as well as aspartic proteases; its binding mode has not been structurally studied so far. Here, we present the crystal structure of the complex of PDI and the serine protease trypsin at 1.95 Å resolution. The binding mode of PDI is compared with that of other Kunitz inhibitors, and structural variability of their reactive centers is discussed.