Synthesis of the segment (11–23) located in the first tandem repeat of plasma kallikrein: comparative binding studies of this and another segment (328–343) to high-molecular-mass kininogen

The synthesis of porcine plasma kallikrein (pPK) segment (11–23), of sequence Phe-Phe-Arg-Gly-Gly-Asp-Val-Ser-Ala-Met-Tyr-Thr-Pro, present in the first tandem repeat sequence of the regulatory chain of PK, has been accomplished following the peptide fragments (5 + 4 + 4) condensation strategy in solution, as well as by fluorenylmethoxycarbonyl solid-phase chemistry. This and another synthetic PK segment of residues (328–343) present in the fourth tandem repeat sequence [Cys(ACM)-Ser-Leu-Arg-Leu-Ser-Thr-Asp-Gly-Ser-Pro-Thr-Arg-Ile-Thr-Tyr] and synthesized by a solid-phase method, were fully characterized by ¹H nuclear magnetic resonance, fast atom bombardment mass spectrometry, amino acid composition and reversed-phase high-performance liquid chromatography. Proteolysis of these peptides by either rat PK (rPK) or trypsin resulted in cleavages between Arg↓Gly for pPK (11–23) and between Arg↓Leu and Arg↓Ile for rPK (328–343). Kinetic studies revealed that for peptide pPK (11–23), the catalytic efficiency (k_cat/K_m) of rPK is 9-fold higher than that of trypsin, but for the other peptide, rPK (328–343), k_cat/K_m of trypsin is 49-fold higher than that of rPK. The facile cleavage of pPK (11–23) by rPK confirms the Arg¹³↓Gly¹⁴ position as the site of autolytic degradation of PK and also explains its special preference for Phe-Phe-Arg sequence.