Time course of HK-D4 cleavage by Sap1, Sap3, Sap5, and Sap6. HK-D4 (10 μM) was cleaved using 0.2 μM Sap in 50 mM citrate buffer (pH 5.0) at 37°C for the specified time. The reaction was stopped using HCl, and the samples were analyzed using HPLC as specified in Figure 1. The results from representative kinetic experiments are shown. The areas under the peaks of the early, major cleavage products, as well as the kinin-related peptides, are expressed relative to the substrate at the beginning of the reaction.