To determine whether AtSBT1.1 has specificity for the AtPSK4 recognition site, we conducted an alanine scan through fpAtPSK4, substituting one residue at a time for alanine (Figure 5a ). The reactions were carried out in triplicate, and the reaction rates for each of the substitutions were compared to that for the wild-type substrate. The cleavage site sequence was very sensitive to alanine substitutions. Substituting the first arginine in the di-basic residues (P6 position) resulted in a reaction rate that was less than half that of wild-type. The most sensitive positions were P2-P4, which are modestly conserved among the AtPSK precursors.